1Department of Anesthesia and Critical Care, Beth Israel Deaconess Medical Center; and 2Department of Neurobiology and the Program in Neuroscience, Harvard Medical School, Boston, Massachusetts 02115
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ABSTRACT |
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Malick, Amy, Rew M. Strassman, and Rami Burstein. Trigeminohypothalamic and Reticulohypothalamic Tract Neurons in the Upper Cervical Spinal Cord and Caudal Medulla of the Rat. J. Neurophysiol. 84: 2078-2112, 2000. Sensory information that arises in orofacial organs facilitates exploratory, ingestive, and defensive behaviors that are essential to overall fitness and survival. Because the hypothalamus plays an important role in the execution of these behaviors, sensory signals conveyed by the trigeminal nerve must be available to this brain structure. Recent anatomical studies have shown that a large number of neurons in the upper cervical spinal cord and caudal medulla project directly to the hypothalamus. The goal of the present study was to identify the types of information that these neurons carry to the hypothalamus and to map the route of their ascending axonal projections. Single-unit recording and antidromic microstimulation techniques were used to identify 81 hypothalamic-projecting neurons in the caudal medulla and upper cervical (C1) spinal cord that exhibited trigeminal receptive fields. Of the 72 neurons whose locations were identified, 54 were in laminae I-V of the dorsal horn at the level of C1 (n = 22) or nucleus caudalis (Vc, n = 32) and were considered trigeminohypothalamic tract (THT) neurons because these regions are within the main projection territory of trigeminal primary afferent fibers. The remaining 18 neurons were in the adjacent lateral reticular formation (LRF) and were considered reticulohypothalamic tract (RHT) neurons. The receptive fields of THT neurons were restricted to the innervation territory of the trigeminal nerve and included the tongue and lips, cornea, intracranial dura, and vibrissae. Based on their responses to mechanical stimulation of cutaneous or intraoral receptive fields, the majority of THT neurons were classified as nociceptive (38% high-threshold, HT, 42% wide-dynamic-range, WDR), but in comparison to the spinohypothalamic tract (SHT), a relatively high percentage of low-threshold (LT) neurons were also found (20%). Responses to thermal stimuli were found more commonly in WDR than in HT neurons: 75% of HT and 93% of WDR neurons responded to heat, while 16% of HT and 54% of WDR neurons responded to cold. These neurons responded primarily to noxious intensities of thermal stimulation. In contrast, all LT neurons responded to innocuous and noxious intensities of both heat and cold stimuli, a phenomenon that has not been described for other populations of mechanoreceptive LT neurons at spinal or trigeminal levels. In contrast to THT neurons, RHT neurons exhibited large and complex receptive fields, which extended over both orofacial ("trigeminal") and extracephalic ("non-trigeminal") skin areas. Their responses to stimulation of trigeminal receptive fields were greater than their responses to stimulation of non-trigeminal receptive fields, and their responses to innocuous stimuli were induced only when applied to trigeminal receptive fields. As described for SHT axons, the axons of THT and RHT neurons ascended through the contralateral brain stem to the supraoptic decussation (SOD) in the lateral hypothalamus; 57% of them then crossed the midline to reach the ipsilateral hypothalamus. Collateral projections were found in the superior colliculus, substantia nigra, red nucleus, anterior pretectal nucleus, and in the lateral, perifornical, dorsomedial, suprachiasmatic, and supraoptic hypothalamic nuclei. Additional projections (which have not been described previously for SHT neurons) were found rostral to the hypothalamus in the caudate-putamen, globus pallidus, and substantia innominata. The findings that nonnociceptive signals reach the hypothalamus primarily through the direct THT route, whereas nociceptive signals reach the hypothalamus through both the direct THT and the indirect RHT routes suggest that highly prioritized painful signals are transferred in parallel channels to ensure that this critical information reaches the hypothalamus, a brain area that regulates homeostasis and other humoral responses required for the survival of the organism.
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INTRODUCTION |
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Trigeminal sensory information that
arises in orofacial organs facilitates exploratory, ingestive, and
defensive behaviors that are essential to overall fitness and survival
(Dubner et al. 1978; Feindel 1956
;
Geppetti et al. 1988
; Guyton 1971
;
Lund and Dellow 1971
). In the rat, tactile signals that
arise in orofacial organs such as the vibrissae, nose, lips, and tongue
contribute to the perception of subtle external cues that assist in the
execution of feeding-related activities such as chewing, swallowing,
licking, and suckling, while painful sensation that originates in these organs can alert the animal to potential dangers and allow for preservation of structures that subserve additional sensory modalities including vision, olfaction, and audition.
Because organs such as the mouth, nose, and eyes serve functions that
must be performed continuously for survival, pain that originates in
these structures is repeatedly aggravated and is therefore one of the
most commonly cited sources of discomfort in human patients. Injuries
such as facial lacerations (Bakay and Glasauer 1980) and
diseases such as trigeminal neuralgia (Kugelberg and Lindblom
1959
), headache (Olesen et al. 1993
), sinusitis
(Saunte and Soyka 1994
), toothache (Sharav
1994
), and temporomandibular joint pain syndrome (Sessle
and Hu 1991
) are believed to activate trigeminal nociceptors
and consequently second-order trigeminal brain stem nuclear complex
(TBNC) neurons in nucleus caudalis and C1-2. Until now,
however, most studies of TBNC neurons that process nociceptive and
nonnociceptive information that arises in the cornea, nasal mucosa,
tongue, tooth pulp, vibrissae, temporomandibular joint, facial muscles,
and skin focused on local or thalamic projecting neurons that are
likely to play a role in the sensory-discriminative aspect of
trigeminal sensation and pain (reviewed in Malick and Burstein
1998
). The purpose of this study was therefore to characterize TBNC neurons that project to brain areas that regulate behavioral, rather than sensory-discriminative, responses to pain (i.e., to determine what kind of information they convey and how they reach their
targets). Because sensations that originate in orofacial organs often
change hypothalamic-mediated behaviors such as food intake and sleep,
we chose to study how orofacial sensory signals reach the hypothalamus.
Complex hypothalamic-mediated functions are commonly influenced by
sensory and physiological signals arising from the body and cognitive
signals arising from cortical and subcortical brain regions. The
integration of sensory, physiological, and cognitive signals by
hypothalamic neurons that regulate both hormonal secretion and the
activity of brain stem and spinal cord neurons that mediate autonomic
responses could provide a partial answer to the question of how sensory
signals produce endocrine, autonomic, and affective responses. To be in
a position to integrate somatosensory and visceral information with
endocrine and autonomic responses, hypothalamic neurons must receive
somatosensory and visceral inputs. The afferent inputs that the
hypothalamus receives from brain stem nuclei, such as the parabrachial
nuclei (Cechetto et al. 1985; Saper and Loewy
1980
; Slugg and Light 1994
), nucleus of the
solitary tract (Menetrey and Basbaum 1987
;
Ricardo and Koh 1978
), periaqueductal gray (Beitz
1982
; Eberhart et al. 1985
; Lima and
Coimbra 1989
; Liu 1983
), and caudal
ventrolateral medulla (Lima et al. 1991
; Sawchenko and Swanson 1981
), and the identification of
neurons in these nuclei that respond to noxious and innocuous
somatosensory and visceral stimulation (Bernard and Besson
1990
; Kannan et al. 1986
; Pan et al.
1999
; Person 1989
; Zhang et al.
1992
) contributed to the notion that somatosensory signals
reach the hypothalamus through several polysynaptic pathways.
Recent anatomical studies showed that somatosensory and visceral
information can also reach the hypothalamus through monosynaptic pathways that originate in spinal cord and medullary dorsal horn neurons. The electrophysiological studies that followed described the
course of sacral, lumbar, thoracic, and lower cervical
spinohypothalamic tract (SHT) axons that convey to the hypothalamus
sensory signals originating in the perineum and colorectal canal
(Katter et al. 1996a), lower limbs (Burstein et
al. 1991
), abdominal region and bile duct (Zhang et al.
1999b
), upper limbs (Dado et al. 1994a
; Kostarczyk et al. 1997
; Zhang et al.
1995
) and cervical dermatomes (Dado et al.
1994b
). Currently, however, no information is available on
axons of TBNC neurons that carry to the hypothalamus sensory information that originates in orofacial organs innervated by the
trigeminal nerve (Burstein et al. 1990
; Carstens
et al. 1990
; Iwata et al. 1992
; Li et al.
1997
; Malick and Burstein 1998
; Newman et
al. 1996
; Ring and Ganchrow 1983
). To fill this
gap, we sought to identify and physiologically characterize
hypothalamic projection neurons in the caudal medulla and upper
cervical spinal cord using antidromic microstimulation and single-unit
recording techniques. Our hypothesis was that the trigeminohypothalamic
tract (THT) is capable of transferring to the hypothalamus nociceptive
and nonnociceptive information that arises in all orofacial organs innervated by the trigeminal nerve.
During the search for THT neurons, we occasionally encountered neurons
that projected to the hypothalamus and responded to noxious stimulation
of both orofacial and extracephalic receptive fields. Later anatomical
analysis revealed that these neurons were located along the poorly
defined ventromedial border of lamina V, in an area previously
identified as the lateral reticular formation (Nord and Kyler
1968). The input that neurons in the lateral reticular formation (LRF) receive from nociceptive neurons in the spinal cord
(Westlund and Craig 1996
) and their well-documented
projections to the hypothalamus (Cunningham and Sawchenko
1991
; Loewy et al. 1981
; McKellar and
Loewy 1981
; Sawchenko and Swanson 1981
) make them additional candidates to transmit nociceptive information to the
hypothalamus. Because our aim is to understand as completely as
possible how sensory trigeminal information reaches the hypothalamus, these reticulohypothalamic tract (RHT) neurons were also studied.
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METHODS |
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Surgical preparations, neuronal recording, and identification of hypothalamic projecting neurons
Male Sprague-Dawley rats weighing 400-600 g were anesthetized with urethan (1.2 g/kg). A metal tube was inserted into the trachea for artificial ventilation, and the rat was mounted in a stereotaxic apparatus. Core temperature was maintained at 37°C by a feedback-controlled heating pad, and end-tidal CO2 was monitored and kept at 4.0-4.5%. A laminectomy was carried out to expose the first cervical segment of the spinal cord (C1), and portions of the occipital bone were removed to allow complete access to nucleus caudalis (Vc) in the caudal medulla. The dura was retracted, the pia removed, and a pool of warm mineral oil formed over the exposed area. Large portions of the frontal and parietal bones were removed on both sides to allow introduction of stimulating electrodes into the hypothalamus, basal ganglia and midbrain. Rats were then paralyzed with gallamine triethiodide (1 g/kg) and artificially ventilated.
Using stainless steel (8-12 M) or tungsten (4-6 M
)
microelectrodes (FHC), single units were recorded within the dorsal
horn of C1, the medullary dorsal horn of Vc, and
the lateral reticular formation (LRF). To search for neurons that
project to the hypothalamus, one or two monopolar stimulating
electrodes were lowered into the contralateral hypothalamus and
cathodal current pulses were delivered (500 µA, 200 µs, 10 Hz).
When one stimulating electrode was used, it was placed in the
anterior-lateral hypothalamus. When two stimulating electrodes were
used, the second was placed in the dorsal-medial area of the posterior
hypothalamus. After isolating the spikes of an antidromically activated
neuron, the stimulating electrode from which the unit was
antidromically activated was moved systematically through the
hypothalamus (as described in Burstein et al. 1991
;
Dado et al. 1994a
) until a point was found from which a
current of
50 µA was capable of inducing consistent antidromic
spikes in the neuron. Criteria for antidromic activation included
constant latency (total variation
0.2 ms), ability to follow trains
of high-frequency stimuli (>333 impulses/s), and collision of
antidromically induced spikes with those induced orthodromically
(Lipski 1981
). All neurons described in this study were
antidromically activated from at least one point in the hypothalamus by
a current of
50 µA. Locations from which neurons were activated antidromically by currents of
50 µA were defined as low-threshold points. Action potentials were amplified, sent to a window
discriminator, collected by computer, analyzed quantitatively by
Neuro-spike software (Pearson Technical Software), and presented as
peristimulus time histograms (500-ms binwidth).
Receptive-field mapping
Following the identification of neurons that project to the
hypothalamus their cutaneous and intraoral receptive fields were mapped
by applying brief innocuous (vibrissae and hair deflection, air puff,
and brush) and noxious (pressure, pinprick, and pinch) mechanical
stimuli to the nose, vibrissal pad, upper and lower lips, tongue, skin
areas above the eye (ophthalmic) and below the eye (maxillary), on the
ventral surface of the face (mandibular), and on the entire body. An
area was considered outside the neuron's cutaneous receptive field if
no stimulus was capable of producing a response in 50% of the
trials. Neurons exhibiting restricted orofacial receptive fields were
classified as "trigeminal" neurons (i.e., trigeminohypothalamic
tract units) and neurons exhibiting both orofacial and extracephalic
receptive fields (e.g., abdomen, limbs, tail) were classified as
"non-trigeminal" neurons (i.e., reticulohypothalamic tract units).
Noncutaneous receptive fields such as the cornea and intracranial dura
were mapped by sliding a brush over these organs and by indenting them
with calibrated von Frey hairs.
Physiological characterization
Neurons were then physiologically characterized according to
their responsiveness to a series of brief (10 s) innocuous and noxious
mechanical stimuli applied to the most sensitive portion of their
cutaneous receptive field. Innocuous stimuli consisted of slowly
passing a soft bristled brush across the cutaneous receptive field and
pressure applied with a loose arterial clip. Noxious stimuli consisted
of pinch with a strong arterial clip and crush with nonserrated
forceps. To avoid inducing prolonged changes in spontaneous neuronal
discharge or response properties, more intense or prolonged stimuli
were not used. Neurons classified as low threshold (LT) responded
maximally or exclusively to innocuous mechanical stimulation. Neurons
classified as wide dynamic range (WDR) responded to brush and also to
noxious mechanical stimulation in a graded fashion. Neurons designated
as high threshold (HT) did not respond to brush but responded to more
intense mechanical stimuli (pressure, pinch, and crush) of their
cutaneous receptive fields (Dado et al. 1994b;
Palecek et al. 1992
). To further characterize the
neurons, their responses to thermal stimulation were determined following the application of thermally conductive paste to the skin. In
most cases, thermal responses were determined by rapidly (10°C/s)
heating (to 39, 41, 46, 50, and 55°C) or cooling (in most cases to
20, 10, and 0°C, in several cases to 30, 25, 20, 15, 10, 5, 0, and
10°C) the skin with a 9 × 9 mm contact thermal stimulator
(Yale University) for 30 s. The data obtained from the rapid-ramp
thermal stimuli were used for the quantitative analyses of the response
magnitude. Thermal responses were also determined by slowly heating
(35-55°C at a rate of 2.4°C/s) or cooling (35-0°C at a rate of
2.0°C/s) the skin (Burstein et al. 1998
) for 10-30 s.
The data obtained from the slow-ramp thermal stimuli were used to
determine response thresholds (Burstein et al. 1998
).
The skin surface was maintained at 35°C during the periods between
stimuli. This period is defined as the interstimulus interval, which
was 180 s. Because we examined only a small number of lamina I
neurons and because not all were examined for their responses to heat,
cold, and mechanical stimuli, we opted not to use the classification of
thermoreceptive-specific (cold), and polymodal nociceptive (HPC)
neurons (Craig and Dostrovsky 1991
; Craig and
Serrano 1994
; Dostrovsky and Craig 1996
;
Han et al. 1998
).
Physiologically characterized units were further classified according
to whether they process sensory information that arises in the
vibrissae, tongue, cornea, intracranial dura, or the entire body. To
identify neurons that respond to vibrissal stimulation, we deflected
individual vibrissae within the neuron's receptive field. The vibrissa
that induced the largest response was then manually deflected in four
orthogonal directions (in 90° increments relative to the horizontal
alignment of the whisker row) at 10-s intervals. Each deflection lasted
5 s. To identify neurons that respond to intraoral stimulation,
the tongue was gently pulled out and exposed to the same mechanical and
thermal stimuli that were applied to the skin. Cornea-sensitive neurons
were identified by gently sliding a brush over the corneal surface;
activating LT A rapidly adapting mechanosensitive receptors
(Giraldez et al. 1979
; MacIver and Tanelian
1993a
,b
). To activate other corneal receptors and nociceptors
(i.e., C-fiber cold receptors, A
high-threshold mechano-heat
nociceptors, and C-fiber chemosensitive receptors) a small portion of
gelfoam dipped in 0.1 M of nicotine (temp = 35°C, pH =7.4) was
laid on top of the corneal surface for 30 s (MacIver and
Tanelian 1993a
,b
; Tanelian and Bisla 1992
), then rinsed with physiological saline. Dura-sensitive neurons were identified by applying single shocks (0.8 ms, 0.5-4.0 mA, 1 Hz) through a bipolar stimulating electrode placed on the dura overlying the ipsilateral transverse sinus (Burstein et al. 1998
).
These stimulus parameters were capable of activating both A
and C
fibers that innervate the dura (Strassman et al. 1996
).
The sinus area from which the lowest current was capable of activating
the neuron was then explored by mechanical stimuli such as dural
indentation with calibrated von Frey hairs (Stoelting, flat and round
tip shape, diameter range = 0.15-0.38 mm) and gentle rubbing with a brush.
When neurons were found to have both orofacial and extracephalic
receptive fields, identical series of mechanical and thermal stimuli
(described above) were applied to each area (on the face, limbs, etc.)
to determine whether the information they process from different skin
regions is qualitatively and/or quantitatively similar. Each series of
stimuli was separated by 3 min. These neurons were classified
separately for their responses to orofacial versus extracephalic stimulation.
Axonal mapping in the midbrain, hypothalamus, and basal ganglia
Once physiological characterization of THT and RHT neurons was
completed, we mapped the course of their axons in the midbrain, hypothalamus, and basal ganglia by using the antidromic
microstimulation mapping technique (for detailed description, see
Burstein et al. 1991; Dado et al.
1994a
,c
; Fields et al. 1995
; Zhang et al.
1995
). To determine the course of the axon, the
hypothalamic-stimulating electrode had to be repositioned. Before
moving this electrode, a second stimulating electrode was inserted into
the contralateral midbrain and placed ~1 mm lateral to the
periaqueductal gray at the level of the superior colliculus. The
position of the midbrain stimulating electrode was adjusted until the
same neuron was activated using a current of
50 µA. To avoid damage
to the parent axon, no attempt was made to lower the current by placing
the electrode closer to the axon. The midbrain stimulating electrode
was used to ensure that the neuron was not lost during the mapping of
its axon when we were unable to activate it from other hypothalamic areas, to recognize the changes that occurred in the amplitude of the
spike during the search, and to confirm that the spike elicited from
the two stimulating electrodes propagated in the same axon. This last
task was achieved by demonstrating that stimulation at the midbrain and
hypothalamus (or any other more rostral point) induced spikes that were
similar in shape, duration and amplitude, and antidromic spikes that
collided with each other when the interspike interval was shorter than
the time required for the spike to travel between the two stimulating electrodes.
To determine whether the axon of the examined neuron terminated in the contralateral hypothalamus, the hypothalamic stimulating electrode was moved as far rostral as the craniotomy allowed and reinserted into the brain in a systematic way that enabled us to determine thresholds for antidromicity at points separated by 200 µm dorsoventrally and 300-500 µm mediolaterally. If the neuron was not activated from any point within the most anterior level, the stimulating electrode was moved 500-1,000 µm posteriorly, and a similar search was made. At each anteroposterior level, the presence of the axon was indicated by a shift in latency of the antidromic spike to a value longer than that recorded from the contralateral midbrain. If a low-threshold point in the contralateral hypothalamus could be surrounded anteriorly, medially, laterally, dorsally, and ventrally by points from which higher currents were required to activate the neuron and if the spikes elicited from that point collided with spikes elicited from the midbrain, the axon was considered to terminate in the contralateral hypothalamus.
To determine whether the axon of the examined neuron crossed the
midline, the stimulating electrode was moved to the ipsilateral side (1 mm from the midline) and repeatedly inserted along the midline
(intervals of 500 µm) from the anterior diencephalon to the midbrain.
In cases in which the neuron was antidromically activated from the
ipsilateral side, the systematic mapping of the axon continued on both
sides of the brain. In cases in which the neuron was not activated from
the ipsilateral side, attempts were made to determine whether the
parent axon issued collateral branches in the hypothalamus. Detailed
description of collateral branches mapping with antidromic stimulation
technique and their limitations are given in our recent paper
(Fields et al. 1995). Briefly; the presence of a branch
was indicated by a shift in latency of the antidromic spike to a value
longer than that of the parent axon at the same anteroposterior level.
The criteria used to confirm that the longer-latency spike was elicited
from a branch of the parent axon were that the position of the
low-threshold point for the putative branch be in one of the
hypothalamic nuclei and at a clear distance from the parent axon in the
supraoptic decussation (where most parent axons are found), and that
the minimum current sufficient to activate the branch be too low to activate the parent axon by current spread.
Anatomical analysis
At the conclusion of each experiment, the recording site and the
low-threshold points for antidromic activation were marked with
electrolytic lesions (anodal DC of 25 µA for 20 s). Only one
neuron was studied in each animal. In cases in which multiple low-threshold points were found, lesions were made at those points from
which a clear shift in latency could be demonstrated. Conduction distances were measured between the recording site and midbrain by
placing the midbrain stimulating electrode over the recording site and
then calculating the differences from the anteroposterior, dorsoventral, and mediolateral stereotaxic coordinates as the shortest
distance between the two points. Similar measurements were made between
the midbrain low-threshold point and each of the hypothalamic
low-threshold points. Rats were perfused with 1% potassium
ferrocyanide in 10% formalin. The brain, brain stem, and upper
cervical spinal cord were removed and postfixed for 5 days, during
which time they were also reacted for Prussian blue stain of ferric
ions. The tissue was cut transversely on a freezing microtome (50 µm)
and examined under dark field illumination, which allowed clear
identification of laminar borders in C1 and Vc.
The tissue was then stained for Nissl substance, and the sections were
reexamined under bright field illumination that shows the cytoarchitectonic organization of the different brain stem nuclei and
dorsal horn laminae. Detailed descriptions of the nuclear (C1, Vc, Vi) and laminar (I-V) borders is given
in our anatomical paper on the THT (Malick and Burstein
1998). Briefly, the rostral border of Vc was defined as the
point at which substantia gelatinosa is "displaced" and becomes
contiguous with the spinal tract of V. We considered Vc to extend 2.0 mm caudal to this point and C1 to extend 1.6 mm
beyond the caudal border of Vc (Falls 1984a
,b
; Gobel et al. 1981
; Jacquin et al. 1986a
).
The transition zone between Vc and C1 usually
coincides with the caudal end of the pyramidal decussation.
Laminar borders in Vc and C1 were determined
using previously described criteria (Gobel et al. 1977;
Jacquin et al. 1990
; Strassman and Vos
1993
). While the identification of the boundaries between
laminae I and II, II and III, and IV and V is relatively straightforward, the precise identification of the inner border of
lamina V is difficult to determine. Since there is no obvious difference between lamina V and the lateral reticular formation, the
medial border of lamina V was defined according to the functional properties of the neurons in this area. As stated in the preceding text, all neurons were classified as either THT or LRF-RHT according to
the location of their cutaneous receptive field. The rationale for this
classification is based on the knowledge that lamina V but not LRF
neurons receive direct input from trigeminal primary afferent fibers
(Arvidsson and Rice 1991
; Clarke and Bowsher
1962
; Jacquin et al. 1982
; Marfurt and
Rajchert 1991
) and that the LRF receives input from nociceptive
dorsal horn neurons located in laminae I and V throughout the length of
the spinal cord (Lima et al. 1991
; Marfurt and
Rajchert 1991
).
Data and statistical analysis
The database consisted of measurements of the number of
spikes/second (response) recorded in C1-THT, Vc-THT, and LRF-RHT
neurons. Data organization and analysis were done on the Prophet System (release 4.1), a national computing resource for life science research
sponsored by the National Institutes of Health, Division of Research
Resources. Response magnitude to each stimulus was calculated by
subtracting the mean ongoing activity occurring before the first
stimulus (10 s for mechanical, 30 s for thermal and chemical) from
the mean firing frequency that occurred throughout the duration of each
stimulus. The means of the measurements were plotted against the
mechanical (brush < pressure < pinch < crush, expressed as scale data 1, 2, 3, 4), heat, and cold stimuli. The resulting distributions were tested for normality using the D'Agostino test (D'Agostino 1986), and their central measures were
computed. Comparison of responses among the respective levels of
mechanical, heat, and cold stimuli were performed using appropriate
multiple sample comparison procedures [Newman-Keuls if the data were
normally distributed (parametric), Kruskal-Wallis if nonparametric].
The means of the measurements were also subjected to trend analysis using the Spearman rank correlation
(rs) for mechanical stimuli and
regression analysis for heat and cold. The responses of LRF-RHT neurons
to mechanical stimulation of their trigeminal and non-trigeminal receptive fields were subjected to unpaired two-sample comparison tests
(unpaired t-test for parametric data, Mann-Whitney rank-sum test for nonparametric data). Neuronal responses to thermal stimuli were analyzed in two ways: during the dynamic phase and during the
static phase. The dynamic phase (heating or cooling ramp) was defined
as the time during which skin temperature is increasing or decreasing
and the static phase as the time during which the temperature was
maintained at constant temperature.
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RESULTS |
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Physiological characterization
IDENTIFICATION OF HYPOTHALAMIC-PROJECTING NEURONS.
Eighty-one neurons (success rate of ~1:3) were antidromically
activated from the contralateral hypothalamus with currents of 50
µA (mean ± SE was 19 ± 11.8 µA). An example of the
localization of a low-threshold point for antidromic activation of a
THT neuron from the contralateral hypothalamus is shown in Fig.
1. In the first track
from which the neuron was antidromically activated (the most medial
track in the hypothalamus), the lowest threshold was 260 µA. After
antidromic thresholds were determined every 200 µm throughout the
track, the electrode was removed and reinserted 300 µm lateral to the
first track. The lowest antidromic threshold in the second track was 70 µA. The electrode was again removed and reinserted 300 µm lateral
to the second track. The lowest antidromic threshold in the third track
was 8 µA. In the next two tracks (made 300 and 600 µm lateral to
the 3rd track), the lowest antidromic thresholds were 32 and 224 µA,
respectively. Since the lowest threshold point in the third track was
surrounded medially, laterally, dorsally, and ventrally by points from
which higher current was required to activate the neuron, it was
considered as the lowest threshold point at this anterior-posterior
level. This point was located in the supraoptic decussation (SOD)
within the lateral hypothalamus (Fig. 1A). Antidromic action
potentials elicited from this and all other low-threshold points in the
hypothalamus for this and all other neurons included in the study
fulfilled the standard criteria for antidromic activation: they
occurred at constant latency, 6.2 ms in this case (Fig.
1B1), collided with orthodromic action potentials elicited
by stimulating the cutaneous receptive field (Fig. 1B2), and
followed a train of high-frequency stimulation (Fig. 1B3).
The recording site of this HT-THT neuron was found in laminae I-II of
Vc (C), and the receptive field was mostly within the
territory of the maxillary branch of the trigeminal nerve
(D).
|
RECORDING SITES. Thirty-two neurons were recorded in Vc, 22 in C1, 18 in LRF, and the locations of 9 neurons were not identified. Photomicrographs of lesions made in laminae I-II and IV-V, and in the LRF are shown in Fig. 2. Reconstructions of the locations of electrolytic lesions marking the recording sites of 72 neurons are illustrated in Fig. 3. Because many lesions were found at the border between laminae II and III and between laminae IV and V, it was difficult to assign each lesion to a particular lamina with certainty. Based on the center of the lesions that were made in Vc and C1, however, it appears that 12 of the neurons were recorded in laminae I-II (20%), 7 in laminae III-IV (15%), and 35 in lamina V (65%). As explained in the preceding text, these 54 neurons were considered trigeminal because their receptive fields were restricted to skin areas innervated by the trigeminal nerve. The lesions of 18 additional recording sites were found in the medullary LRF. Although there is no easy way to differentiate between lamina V and the LRF, neurons assigned to the LRF were found deeper than the lamina V THT neurons and exhibited extracephalic, in addition to orofacial, receptive fields. They were therefore considered non-trigeminal neurons. The recording locations of physiologically characterized neurons in C1 and Vc were distributed as follows: laminae I-II contained 4 HT, 4 WDR, and 1 LT neurons; laminae III-IV contained 1 HT, 3 WDR, and 2 LT neurons; lamina V contained 11 HT, 12 WDR, and 8 LT neurons; and the LRF contained 7 HT, 7 WDR, and 2 LT neurons (LRF classification was based on responses to facial stimulation). Of the unclassified neurons, three were in laminae I-II, one in laminae III-IV, four in lamina V, and two in the LRF.
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RECEPTIVE FIELDS. Twenty of the 22 C1-THT neurons had restricted ipsilateral orofacial receptive fields, and 2 receptive fields were restricted to the ipsilateral neck (Fig. 4A). As shown in the figure, all laminae I and II C1-THT neurons exhibited small to medium receptive fields that extended over facial skin areas innervated by one or two branches of the trigeminal nerve, while many laminae III-V neurons exhibited medium to large receptive fields that extended over facial skin areas innervated by two to three branches of the trigeminal nerve. Similar receptive fields were mapped for the 32 Vc-THT neurons (26 of which are shown in Fig. 4B); laminae I-II neurons had primarily small receptive fields, whereas those located in deeper laminae exhibited large receptive fields as well. In general, most HT-THT neurons had small or medium receptive fields, and most WDR-THT neurons had medium or large receptive fields. This tendency, however, was influenced by their location in the different laminae; both HT and WDR neurons had smaller receptive fields if they were recorded in laminae I-II and larger receptive fields if they were recorded in laminae III-V.
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HIGH THRESHOLD NEURONS. Twenty-four of the 64 (38%) physiologically characterized neurons responded exclusively to noxious mechanical stimuli and were therefore classified as HT. Examples of the responses of two HT-THT neurons are illustrated in Fig. 5. The neuron on the left (Fig. 5A) was recorded in the most dorsomedial portion of lamina V, exhibited a mandibular/maxillary receptive field, and was antidromically activated from the contralateral hypothalamus. It responded to noxious but not innocuous mechanical stimuli (Fig. 5C). The neuron on the right (Fig. 5B) was recorded in the most ventrolateral portion of lamina I, exhibited an ophthalmic receptive field, and was antidromically activated from the lateral hypothalamus. It also responded exclusively to the noxious mechanical stimuli (Fig. 5D). The mechanical response profiles of 23 HT neurons are illustrated in Fig. 5E. Their mean (± SE) firing rates to brush, pressure, pinch and crush were 0.2 ± 0.1, 5.0 ± 1.6, 29.0 ± 5.5, and 36.0 ± 4.8 spikes/s, respectively.
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WIDE-DYNAMIC RANGE NEURONS. Twenty-seven of the 64 (42%) physiologically characterized neurons responded to innocuous and noxious stimuli in a graded fashion and were therefore classified as WDR. Examples of the responses of two WDR-THT neurons are illustrated in Fig. 6. The neuron on the left (Fig. 6A) was recorded in the dorsomedial portion of lamina V, exhibited a large mandibular/maxillary/ophthalmic receptive field, and was antidromically activated from the contralateral hypothalamus. It responded most vigorously to innocuous and noxious mechanical stimulation of its mandibular receptive field; stimulation of its maxillary and ophthalmic receptive field produced smaller responses (Fig. 6C). The neuron on the right (Fig. 6B) was recorded in the most ventrolateral portion of lamina V, exhibited a large ophthalmic/maxillary/mandibular receptive field, and was antidromically activated from the contralateral hypothalamus. It responded most vigorously to innocuous and noxious mechanical stimulation of its ophthalmic receptive field; stimulation of its maxillary and mandibular receptive field produced smaller responses (Fig. 6D). The mechanical response profiles of 27 WDR neurons are illustrated in Fig. 6E. Their mean (± SE) firing rate to brush, pressure, pinch, and crush were 8.3 ± 1.2, 26.6 ± 3.6, 40.2 ± 3.8, and 40.5 ± 2.7 spikes/s, respectively.
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LOW THRESHOLD (VIBRISSA-SENSITIVE) NEURONS. Thirteen of the 64 (20%) physiologically characterized neurons responded more vigorously to innocuous than to noxious stimuli and were therefore classified as LT. Most of these LT neurons responded to deflection of a single hair follicle or vibrissa. Examples of the responses of a vibrissa-sensitive LT-THT neuron are illustrated in Fig. 7. This neuron was recorded in the ventrolateral portion of laminae III-IV, exhibited a small receptive field, and was antidromically activated from the contralateral hypothalamus and ventromedial posterior thalamic nucleus (Fig. 7A). It responded maximally to the deflection of a single vibrissa in all four directions (Fig. 7B) and to brushing its receptive field (not shown). The mechanical response profiles of 13 LT neurons are illustrated in Fig. 7C. Their mean (± SE) firing rates to brush, pressure, pinch, and crush were 34.0 ± 3.0, 21.3 ± 5.5, 22.0 ± 4.7, and 24.5 ± 5.0 spikes/s, respectively.
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RESPONSES TO THERMAL STIMULI. Heat. Innocuous and noxious heat stimuli were applied to the receptive fields of 29 neurons. Twenty-seven of these (93%) responded incrementally to graded increases in heat stimuli. Of the 27 heat-sensitive neurons, 5, 3, and 9 were recorded in laminae I-II, III-IV, and V of C1-Vc, respectively, and 10 were recorded in the LRF. Figure 8 illustrates two different response types to heat stimuli: at left; a "static" response, defined as maximal discharge during the steady-state phase of the stimulus, and at right; a "dynamic" response, defined as maximal discharge during the heating ramp.
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ORAL-SENSITIVE NEURONS. Twenty-one neurons responded to mechanical stimulation of the oral mucosa, tongue, or lips. They were classified as HT in 9 cases, WDR in 9 cases, and LT in 3 cases. The majority of oral-sensitive THT neurons were recorded in the dorsomedial third of laminae V (11 units) and III-IV (2 units) of C1 (6 units), and Vc (7 units). The other seven oral-sensitive neurons were recorded in the LRF, and the recording site of one neuron was not found. The cutaneous receptive fields of the THT neurons varied; they extended over small mandibular or maxillary areas in three (23%) cases, mandibular and maxillary areas in 4 (31%) cases, and the entire face in six (46%) cases. Regardless of the cutaneous receptive field size, maximal neuronal responses were most commonly induced by stimulating intraoral structures. An example of an oral-sensitive HT-THT neuron is illustrated in Fig. 10 (left). This hypothalamic projecting neuron was recorded in the most dorsomedial portion of lamina V and exhibited an oral receptive field that included the tongue, hard palate, and the upper lip (A). It responded more vigorously to noxious mechanical stimuli of the hard palate than the tongue (B). The mechanical response profiles of 19 oral-sensitive neurons are illustrated in C. Their mean (± SE) firing rates to brush, pressure, pinch, and crush were 4.7 ± 1.6, 12.2 ± 3.0, 30.6 ± 4.6, and 37.2 ± 4.2 spikes/s, respectively. The heat response profiles of eight oral-sensitive neurons are illustrated in D. Their mean (± SE) firing rates to 39, 41, 46, 50, and 55°C were 0.1 ± 0.1, 0.3 ± 0.3, 4.1 ± 1.5, 17.9 ± 6.3, and 27.6 ± 8.6 spikes/s, respectively. About 15% of the oral-sensitive THT neurons also responded to innocuous cold stimuli (data not shown).
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CORNEA-SENSITIVE NEURONS. Thirteen neurons responded to mechanical stimulation of the cornea. They were classified as HT in four cases, WDR in seven cases, and LT in two cases. All THT neurons were recorded in lamina V at the level of caudal Vc (5 units) and rostral C1 (4 units). Of the remaining four neurons, three were recorded in the LRF, and the recording location of one unit was not identified. Although no attempt was made to map the corneal receptive fields of these neurons, they seemed to respond to stimulation of all four corneal quadrants. In all but one case, the cutaneous receptive field extended over the periorbital skin; they included the ophthalmic and maxillary skin in all cases, and the mandibular skin in less than half of the cases. An example of a cornea-sensitive THT neuron is illustrated in Fig. 10 (middle). This hypothalamic projecting neuron was recorded in the most lateral region of lamina V, and its receptive field included the cornea and the periorbital skin (A). It responded preferentially to noxious mechanical stimuli of the ophthalmic skin and most vigorously to mechanical and chemical (nicotine) stimulation of the cornea (B). The mechanical response profiles of 10 cornea-sensitive neurons are illustrated in C. Their mean firing rates to brush, pressure, pinch, and crush were 10.8 ± 5.8, 31.0 ± 10.0, 41.7 ± 7.2, and 39.6 ± 6.5 spikes/s, respectively. The heat response profiles of nine cornea-sensitive neurons are illustrated in D. Their mean firing rates to 39, 41, 46, 50, and 55°C were 0.1 ± 0.1, 0.8 ± 0.5, 3.4 ± 1.5, 7.8 ± 1.6, and 15.1 ± 3.5 spikes/s, respectively. In five cases, 1 M nicotine was applied to the cornea for 30 s. The neuronal responses in all cases were vigorous.
DURA-SENSITIVE NEURONS. Ten THT neurons responded to electrical and mechanical stimulation of the dura mater overlying the transverse sinus. The majority (8 units) were recorded in the most lateral region of lamina V at the level of Vc, and only two neurons were recorded in laminae I-III or in C1. The dural receptive fields of these neurons were usually small and restricted to the transverse or superior sagittal sinuses. All dura-sensitive neurons also exhibited cutaneous receptive fields (which varied in size). Regardless of the cutaneous receptive field size, maximal neuronal responses were most commonly elicited from the periorbital skin region. Eight of the 10 dura-sensitive neurons were physiologically characterized based on their responses to cutaneous stimulation; all were classified as WDR. An example of a dura-sensitive WDR-THT neuron is illustrated in Fig. 10 (right). This hypothalamic projecting neuron (A1) was recorded in the most lateral region of lamina V (A2). It exhibited ophthalmic receptive fields on the skin (A3) and the dura (A4). It responded preferentially to noxious mechanical stimulation of the ophthalmic skin and to dural brushing (B). The mechanical response profiles of eight dura-sensitive neurons are illustrated in C. Their mean firing rate to brush, pressure, pinch, and crush were 9.0 ± 2.4, 38.1 ± 7.3, 50.6 ± 7.4, and 43.1 ± 5.7 spikes/s, respectively. The heat response profiles of four dura-sensitive neurons are illustrated in D. Their mean firing rates to 39, 41, 46, 50, and 55°C were 4.0 ± 3.0, 5.8 ± 2.5, 16.0 ± 4.1, 33.8 ± 7.3, and 33.0 ± 1.0 spikes/s, respectively.
LRF-RHT NEURONS. Eighteen hypothalamic projecting neurons were recorded in the LRF. Their recording sites were in general more medial and slightly more ventral than the recording sites of the THT neurons in the ventrolateral area of lamina V (Fig. 3). Their most distinct property was their cutaneous receptive fields. They included orofacial and extracephalic skin areas in all cases (Fig. 4). In 11 cases, receptive fields extended over the entire body. An example of a HT LRF-RHT neuron is illustrated in Fig. 11. This hypothalamic projecting neuron exhibited a whole body receptive field. Like most LRF-RHT neurons in this study, it responded more vigorously to mechanical stimulation of ipsilateral orofacial organs (i.e., tongue, cornea, skin) compared with mechanical stimulation of contralateral orofacial organs or any of the extracephalic regions (i.e., paws and tail). Figure 12 illustrates the responses of all 18 LRF neurons to mechanical stimulation of their facial (i.e., trigeminal, Fig. 12A) and extracephalic (i.e., non-trigeminal, Fig. 12B) receptive fields. The means (±95% confidence interval) of the responses to brush, pressure, pinch, and crush are shown in Fig. 12C. Responses induced by trigeminal receptive fields stimuli were significantly greater than the responses to the respective stimuli of the non-trigeminal receptive fields (P values given in the right column of the table). These findings indicate that LRF neurons responded to brush and pressure of their trigeminal but not non-trigeminal receptive fields and that pinch and crush induced larger responses when applied to their trigeminal than to non-trigeminal receptive field.
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COMPARISONS OF THE RESPONSE PROFILES OF C1-THT, VC-THT, AND LRF-RHT NEURONS. The response profiles of C1-THT, Vc-THT, and LRF-RHT were compared. These data are presented in Fig. 13, where the response profiles of all HT, WDR, and LT neurons recorded in each of the indicated areas were grouped. Within all three areas, responses to pressure were significantly larger than responses to brush, and responses to pinch were significantly larger than responses to pressure. Among the three groups, the magnitude of the responses to brush (e.g., C1-THT brush vs. Vc-THT brush vs. LRF-RHT brush), pressure, pinch, and crush were not different (Fig. 13A). The mean ± 95% confidence interval of the responses are shown in the tables on the right. The trends of increased response magnitudes with increased stimulus intensity were also significant for all three groups (P values shown in the row marked "correlation").
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Axonal mapping
ANTIDROMIC MAPPING IN THE CONTRALATERAL MIDBRAIN AND DIENCEPHALON.
Anatomical (Burstein et al. 1987; Cliffer et al.
1991
) and physiological (Burstein et al. 1991
;
Dado et al. 1994a
; Katter et al. 1996a
)
studies indicated that most spinohypothalamic tract axons reach the
hypothalamus through the posterior thalamus and the supraoptic
decussation. To determine whether THT and RHT neurons also share this
anatomical approach, we attempted to map their axonal routes between
the midbrain and the hypothalamus. In 72 cases (54 THT and 18 RHT),
neurons that were antidromically activated from the contralateral
hypothalamus were also antidromically activated with currents of
50
µA from the contralateral midbrain and posterior diencephalon. Figure
14 illustrates an
experiment in which a neuron was antidromically activated from
low-threshold points in the contralateral midbrain, contralateral
caudal diencephalon, and contralateral hypothalamus. The neuron was
initially activated antidromically from a lowest threshold point in the
contralateral lateral hypothalamus (point a). The lesion marking the
location of this point was found just medial to the optic tract, within the supraoptic decussation. The antidromic latency from this point was
2.7 ms (Fig. 14C), and the minimum current required to
activate the neuron was 18 µA. When the stimulating electrode was
moved in the medial, lateral, dorsal, or ventral directions, higher currents were required to activate the neuron antidromically. Prior to
the removal of the first stimulating electrode from the lowest
threshold point in the hypothalamus, a second stimulating electrode was
used to search for a low-threshold point for antidromic activation of
that same neuron from the midbrain (point d). This point was found
between the periaqueductal gray and the superior colliculus (Fig.
14B). The antidromic latency from this point was 1.4 ms
(Fig. 14C), and the minimum current required to activate the
neuron was 11 µA. The first stimulating electrode was then moved 1.5 mm posteriorly, and 11 electrode penetrations were made across the
mediolateral extent of the contralateral posterior hypothalamus and
thalamus. At this level, the low-threshold point was also located
between the internal capsule and the optic tract (point b). The
antidromic latency from this point was 2.0 ms, and the minimum current
required to activate the neuron was 14 µA. At the level of the
posterior commissure (3.5 mm posterior to point a), eight electrode
penetrations were made contralaterally. At this level, the lowest
threshold point for antidromic activation was located in the substantia
nigra pars compacta (point c; 1.6 ms, 9 µA). That the same neuron was
antidromically activated from each of the lowest threshold points
(a-c), was confirmed by colliding the antidromic spikes evoked at each
of these points with the antidromic spikes evoked from the second
stimulating electrode in the midbrain (Fig. 14C, d-a, d-b,
and d-c). Based on the recorded latencies from each point and their
distances from the recording site, we estimated that the conduction
velocity of this axon was 6.2 m/s to point a, 7.7 ms to point b, and
8.3 ms to point c.
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ANTIDROMIC MAPPING IN THE CONTRALATERAL AND IPSILATERAL
HYPOTHALAMUS.
Fourteen THT and 7 RHT neurons that were initially activated
antidromically from the contralateral hypothalamus were also tested for
their projections to the ipsilateral hypothalamus. Eight THT (57%) and
4 RHT (57%) neurons were antidromically activated with currents of
30 µA from low-threshold points in the ipsilateral hypothalamus (it
was not within the scope of this study to determine how far caudal
these axons descend). Figure 15
illustrates an example of a case in which the neuron
was antidromically activated from low-threshold points in the
contralateral midbrain, contralateral hypothalamus, and ipsilateral
hypothalamus. The neuron was initially activated from a single
low-threshold point located 1.3 mm caudal to Bregma, within the
contralateral hypothalamus (point f). All antidromic spikes induced
from the contralateral hypothalamus at this level reached the neuron
within 3.3 ms. Following the identification of a low-threshold point in
the midbrain (point a), the stimulating electrode was moved 1 mm
anteriorly (0.3 mm caudal to Bregma), and nine individual electrode
penetrations were made across the mediolateral extent of the preoptic
area, bilaterally. The neuron could not be activated antidromically from any one of the 180 tested points at this level, suggesting that
the axon does not project more anteriorly. The electrode was therefore
moved back to the anterior hypothalamus (1.3 mm caudal to Bregma), and
another set of electrode penetrations was made in the ipsilateral
hypothalamus. A low-threshold point for antidromic activation was
located in the SOD (point g), suggesting that the axon crossed the
midline within the optic chiasm. All antidromic spikes induced from the
ipsilateral hypothalamus at this level needed the same time to reach
the neuron (4.2 ms). The stimulating electrode was repositioned, and 12 individual electrode penetrations were made at the level of the
ventromedial hypothalamic nucleus (2.3 mm caudal to Bregma). At this
plane (on the ipsilateral side), two low-threshold points were found. One was located in the optic tract and the other in the lateral hypothalamus, just ventral to zona incerta (ZI). The antidromic spikes
induced from these low-threshold points differed in their latencies;
the spike induced from the optic tract (point h) reached the neuron in
4.6 ms, and the spike induced from the lateral hypothalamus needed 6.0 ms (point i). Because these low-threshold points were separated by
points from which higher currents were required to induce antidromic
spikes of similar latencies and because the long latencies could not be
induced from point h nor the short latencies from point i, we concluded
that the parent axon in the ipsilateral optic tract issued a collateral
branch in the lateral hypothalamus. When the stimulating electrode was
moved back to the contralateral side, additional low-threshold points
were found in the contralateral SOD (point d) and contralateral lateral
hypothalamus (point e). Since the antidromic spikes induced from these
low-threshold points differed (2.8 ms from point d, and 4.4 ms from
point e), we again concluded that the parent axon in the contralateral
SOD issued a collateral branch in the lateral hypothalamus. By tracking the axon backward on the contralateral side, it was also possible to
activate this neuron antidromically from low-threshold points in the
subthalamic nucleus (point c) and the central tegmental field [point
b, deep mesencephalic nucleus (DPME)] of the midbrain.
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ANTIDROMIC MAPPING IN THE BASAL GANGLIA.
Previous anatomical studies (Burstein and Giesler 1989;
Burstein and Potrebic 1993
; Burstein et al.
1987
; Cliffer et al. 1991
; Malick and
Burstein 1998
; Yasui et al. 1987
) indicated that
some spinal cord neurons, including those located in
C1 and Vc, project to forebrain areas positioned
anterior to the hypothalamus (i.e., nucleus accumbens, globus pallidus,
substantia innominata, septal nuclei, bed nucleus stria terminalis).
Although systematic attempts were not made to explore these anterior
areas thoroughly, we occasionally (3 cases) activated neurons in
C1 and LRF from forebrain nuclei such as the
caudate-putamen, globus pallidus, ventral pallidum, and the substantia
innominata. Figure 16
illustrates antidromic activation of a C1-THT
neuron from several basal ganglia regions. This lamina V-WDR neuron was
initially activated antidromically from the supraoptic nucleus in the
hypothalamus (point g). In addition, it was also possible to activate
this neuron antidromically from the contralateral basal ganglia and
midbrain (Fig. 16, A and B). At the level of the
posterior commissure, only one low-threshold point was found (point k,
3.0 ms). This point was just dorsal to the medial lemniscus, within the
deep mesencephalic/central tegmental field. At the two levels of the
diencephalon (1.7 and 0.7 mm caudal to Bregma), several low-threshold
points were found (Fig. 16B). At the level of
mid-hypothalamus (1.7 mm caudal to Bregma), one low-threshold point was
located in the supraoptic decussation (point j, 4.1 ms), and the other
two were located in the internal capsule (point h, 3.8 ms)
and globus pallidus (point i, 8.0 ms). At the level of the rostral
hypothalamus (0.7 mm caudal to Bregma), one low-threshold point was
located in the supraoptic nucleus (point g, 4.2 ms), and the others in
the caudate-putamen (point c, 4.4 ms), substantia innominata (point d,
5.2 ms), and globus pallidus (point e, 6.6 ms; point f, 7.0 ms). At the
level of the anterior commissure (0.3 mm anterior to Bregma), only one low-threshold point was found (point b, 4.6 ms). This point was located
in the most ventral and lateral area of the caudate-putamen. The most
anterior low-threshold point from which the neuron was antidromically
activated was found at the level of nucleus accumbens (1.3 mm anterior
to Bregma). This point (a) was located just lateral to nucleus
accumbens, within the caudate-putamen. Based on the latencies induced
from low-threshold, points a, b, c g, h, j, and k, and their
distances from the recording site, it seems that the parent axon may
have bifurcated in two directions between the midbrain and the
hypothalamus. One branch seems to approach the supraoptic nucleus
through the supraoptic decussation (points k, j, and g), while the
other seems to approach the ventrolateral caudate-putamen region
through the internal capsule (points h, c, b, and a). In addition,
since the latencies induced from points d, e, f, and i were longer than
the latencies induced from more anterior levels, it is possible that
small collateral branches were activated at these points. The two
smaller branches (h to i and c to d, e and f) may have
emanated from the branch that headed to the caudate-putamen.
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ANTIDROMIC ACTIVATION SITES OF THT AND LRF-RHT NEURONS. Figure 17 illustrates the locations of 197 low-threshold points for antidromic activation of 54 THT (A) and 18 LRF (B) neurons. Regardless of the location of the recording site (A and B) or the physiological classification (C) of the neurons, all axons seemed to follow the same path. At the level of the red nucleus, in the rostral midbrain, 32 low-threshold points for antidromic activation of the ascending axons of 15 THT and 12 RHT neurons were found in a poorly defined area between the central gray, medial lemniscus, substantia nigra, and the superior colliculus. Although uncommon, individual axons could also be found in the ventral region of the superior colliculus (SC), the substantia nigra pars compacta (SNc), and the red nucleus. At the level of the posterior commissure, most axons (of both populations) had the tendency to shift ventrally, toward the cerebral peduncle. In the 13 experiments (i.e., individual neurons) in which the location of the axon was determined at this level, five low-threshold points were found in the posterior thalamus, three in the medial lemniscus, seven in the sub-thalamus, and three in the cerebral peduncle. Occasionally low-threshold points were found in the substantia nigra and the anterior pretectal nucleus (APT). Rostral to the posterior commissure, within the caudal diencephalon, 73 low-threshold points for antidromic activation of the ascending axons of 36 trigeminal and 18 non-trigeminal neurons were found in the internal capsule (6), on the border between the internal capsule and optic tract (7), supraoptic decussation (41), lateral hypothalamus (13), perifornical area (2), dorsomedial hypothalamus (1), zona incerta (2), and ventroposterior medial thalamic nucleus (1). In the rostral diencephalon, the ascending axons of 15 THT and 17 RHT neurons were activated antidromically from 44 low-threshold points that were located in the supraoptic decussation (29), internal capsule (2), supraoptic nucleus (3), suprachiasmatic nucleus (1), lateral hypothalamus (2), globus pallidus (3), caudate putamen (1), substantia innominata (2), and the basal nucleus of Myenert (1). Anterior to the hypothalamus, low-threshold points were found in the ventrolateral area of caudate putamen (2) and in the globus pallidus (1). On the ipsilateral side, 27 low-threshold points for antidromic activation of nine THT and four RHT neurons were found between the anterior hypothalamus and the midbrain. These points were located in the optic chiasm (3), suprachiasmatic nucleus (2), supraoptic decussation (9), optic tract (4), lateral hypothalamus (4), internal capsule (1), cerebral peduncle (2), and medial to the medial geniculate nucleus (2). Photomicrographs of lesions made at low-threshold points in the basal ganglia, hypothalamus, and midbrain are shown in Fig. 2.
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CONDUCTION VELOCITIES. In 58 cases (38 THT and 20 RHT), conduction velocities (CVs) were determined at several points along the axonal path (Fig. 18A). The estimated mean CV between the recording sites in C1-Vc or LRF and the low-threshold points in the caudal hypothalamus was 7.3 ± 0.3 m/s, with a range of 1.2-15.0 m/s (Fig. 18I). The estimated CVs between the recording sites and the midbrain and between the midbrain and caudal hypothalamus were 8.3 ± 0.4 and 7.2 ± 0.7 m/s, respectively (Fig. 18, II and III). Between the caudal and rostral hypothalamus, conduction of THT and RHT axons averaged 3.4 ± 1.4 and 6.1 ± 2.1 m/s, respectively (Fig. 18, IV and V). These data indicate that the axons of THT but not RHT neurons slowed by 50% as they passed through the hypothalamus (P = 0.001; t test); pointing to the possibility of collateral branches.
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CURRENT SPREAD.
The effective spread of the antidromic stimulation currents in the
hypothalamus was estimated by examining the relationship between
stimulus intensity and distance from the axon (as defined by the lowest
threshold point for antidromic activation). The plots in Fig. 18,
I and II, summarize threshold readings from 66 electrode penetrations containing lowest threshold points (i.e., 50
µA) in the hypothalamus. As indicated by the plots, there was a
direct relationship between the distance from the axon and the
threshold for antidromic activation. At 200, 400, 600, and 800 µm
from the axon, the mean effective current spread was 40, 105, 197, and
225 µA. The threshold for antidromic activation of 90% of the
neurons described in this study was
30 µA. Although the mean
effective spread of current
30 µA was smaller than 200 µm (Fig.
18II), it occasionally (2/66) reached as far as 400 µm and
rarely (1/66) beyond this distance (Fig. 18I). Throughout
the study, a current of 500 µA was used in the initial search for hypothalamic projection neurons. The maximum effective spread of this
current was only 1,500 µm (Fig. 18I). These data show that the current-distance relationship within the hypothalamus depends on
the intensity of the current; low currents can effectively activate
axons at distances that suggest a spread of 10-20 µm/µA and high
currents at distances that suggest a spread of 3-5 µm/µA.
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DISCUSSION |
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This study describes the response properties of two ascending somatosensory pathways that convey primarily nociceptive information to the hypothalamus: the trigeminohypothalamic tract, which conveys sensory signals that arise in facial skin, cornea, vibrissae, oral mucosa, and intracranial dura, and the reticulohypothalamic tract, which relays sensory signals from the entire body via the spinal cord and TBNC. To our knowledge, this is the first evidence that sensory information that originates in these orofacial organs reaches the hypothalamus directly. It is also the first study to document the kind of sensory information that RHT neurons in the LRF carry. We consider the THT a direct pathway because it receives direct input from trigeminal primary afferent fibers. We consider the LRF-RHT an indirect pathway because it does not receive direct input from peripheral fibers, rather the input that drives its neurons originates in laminae I and V throughout the length of the spinal and medullary dorsal horn. The findings that nociceptive signals that arise in orofacial organs reach the hypothalamus through both the THT and the RHT suggest that highly prioritized signals regarding orofacial pain are transferred in parallel channels to ensure that this critical information reaches the hypothalamus; a brain area that regulates homeostasis and other humoral responses required for the survival of the organism.
Although the properties of the THT are broadly similar to those described previously for the SHT, a number of fundamentally new findings have emerged from the present study: 1) neurons classified as LT by their response to mechanical stimulation also respond to innocuous and noxious intensities of thermal stimulation. Their markedly different stimulus-response curves during the static and dynamic phases of the thermal stimuli suggest a novel pattern of convergent inputs from thermosensitive and nociceptive primary afferent neurons (see following text). Such thermal responses have not been described for other populations of LT neurons at spinal or trigeminal levels. 2) Somatotopy is expressed in some lamina V WDR neurons as a gradation of sensitivity within a large receptive field such that the most sensitive part of the receptive field corresponds to the neuron's position along the dorsomedial-to-ventrolateral axis. 3) Dorsal horn neurons that project directly to the forebrain, which have been described previously only in anatomical studies, have been physiologically characterized and shown to convey nociceptive information. It has further been shown that the axons of these neurons ascend by a different route than SHT/THT neurons, through the internal capsule rather than through the supraoptic decussation. 4) Neurons in lamina V of Vc can be distinguished physiologically from neurons in the adjacent LRF in that lamina V neurons have trigeminal-only receptive fields while LRF receptive fields include both trigeminal and extra-trigeminal regions.
Responses to cutaneous orofacial stimuli
Although the response properties of THT and RHT neurons to mechanical and thermal stimulation of the orofacial skin appear similar, they differed in their anatomical locations and receptive field properties, suggesting that different inputs govern their responses. They will therefore be discussed separately.
Thirty-eight percent of all THT neurons were classified as HT. They
responded exclusively to noxious mechanical and thermal stimulation and
were capable of reliably encoding the intensity of the noxious
stimulus. Based on the trend in their responses to pressure, pinch, and
crush (but not to brush) and to 46, 50, and 55°C (but not to 39 and
41°C), it is reasonable to propose that their peripheral inputs
originate in cutaneous nociceptors that innervate the hairy skin. In
primates, these nociceptors include type I and II
A-mechanoheat (AMH) and C-mechanoheat (CMH) fibers; AMH and CMH fibers
are also termed HT mechanoreceptors and C polymodal nociceptors,
respectively (Bessou and Perl 1969; Perl
1969
). The early burst of activity, the slow adaptation rate, the monotonic increase in responses to the increased intensities of
both mechanical and thermal stimuli, and the plateau in the responses
to the most intense (heat) stimuli suggest a CMH input (Garell
et al. 1996
; LaMotte and Campbell 1978
;
Meyer and Campbell 1981
), while the graded responses to
the increased intensities of the mechanical stimuli (i.e., the lack of
plateau), and the relatively high heat threshold (47.4°C) point to
type II AMH input (Treede et al. 1995
).
Type II AMH nociceptors respond to lower heat stimuli
(median threshold 46°C) than type I AMH (50°C)
(Treede et al. 1995
), and higher heat stimuli than CMH
nociceptors (43-44°C) (LaMotte and Campbell 1978
),
40-41°C (Tillman et al. 1995a
,b
; Weidner et
al. 1999
). The "lack of" or poor responses to cold stimuli suggest that THT-HT neurons receive most of their peripheral inputs from mechanoheat nociceptors that do not usually respond to cold
stimuli (Simone and Kajander 1997
).
Forty-two percent of all THT neurons were classified as WDR. They
responded preferentially to noxious mechanical stimuli and almost
exclusively to noxious heat stimuli. Their responses to innocuous
mechanical stimuli, and the graded fashion of their responses to
noxious stimuli suggest that the peripheral inputs they receive
originate in both mechanoreceptors and nociceptors. Clues to the source
of their nociceptive input are provided by their different responses to
heat. WDR neurons responded incrementally to the dynamic (i.e., reached
their peak during the dynamic phase of the heat stimulus) and static
(i.e., reached their peak during the later static phase of the heat
stimulus) phases of the heat stimulus. The incremented response to the
static phase of the increased heat stimulus (between 45 and 55°C) and
the heat threshold (43.6°C) suggest an input from CMH (LaMotte
and Campbell 1978) and potentially from type I AMH
(Treede et al. 1995
), while the graded responses to the
dynamic phase could be mediated by signals that originate in type
II AMH (Sumino et al. 1973
; Treede et al. 1995
) and potentially CMH (Meyer and Campbell
1981
) nociceptors. The incremented responses to the static
phase of the increased cold stimuli (between 20 and 0°C) suggest that
THT-WDR neurons can encode the intensity of the noxious cold stimuli
and that their so-called static response to cold is mediated by
nociceptors. Because the responses of WDR neurons to thermal stimuli of
35-20°C were not examined routinely, we cannot rule out or provide
sufficient evidence for a potential influence of cold receptors on the
initiation of their response to cold.
Twenty percent of all THT neurons were classified as LT. Their
responses to the dynamic innocuous mechanical stimuli (brush) were
significantly larger than their responses to the noxious mechanical
stimuli (P < 0.05). Traditionally, these response
profiles are believed to be mediated by the activation of LT
mechanoreceptors (LTM) that provide information about texture and shape
(reviewed in Johnson 1983). Most LT neurons also
responded to deflection of a single vibrissa or hair follicle; stimuli
that are considerably more gentle than brushing the skin. In the rat,
several classes of rapidly and slowly adapting mechanoreceptors that
innervate the vibrissae, guard hairs and F-line upper lip hair are
believed to provide the sensory information required for recognition of objects in the environment (Jacquin et al. 1986a
,b
).
Based on the mechanical response properties of LT neurons, we must
conclude that a part of their input originates in LT mechanoreceptors
that innervate the hairy skin. We are somewhat puzzled, however, by the
thermal response properties of these THT-LT neurons. As shown in Figs.
8 and 9, the responses of LT neurons increased incrementally during the
steady-state phase of the noxious heat and cold stimuli. Since
nociceptors are the only receptors that respond in a graded fashion to
increased intensities of noxious thermal stimuli, we have interpreted
these data as suggesting that the static responses of THT-LT neurons
are influenced by inputs that originate in nociceptors. A possible site
of interaction between primary afferent nociceptors and second-order LT
neurons is lamina V of the dorsal horn, as this lamina receives direct
input from cutaneous nociceptors and contained most (8/11) LT-THT
neurons. Based on the mechanical response profile, however, it is not
reasonable to propose that their inputs originate in cutaneous CMH and
AMH nociceptors. Rather, an input from nociceptors that are heat and/or
cold sensitive, but mechanically insensitive is proposed. Heat
nociceptors have been found in both humans and animals (Baumann
et al. 1991
; Georgopoulos 1976
; Treede et
al. 1998
; Weidner et al. 1999
; Welk et
al. 1984
). In humans, they constitute a class of C-fiber
nociceptors that is clearly distinct from CMH nociceptors; they are
mechanically insensitive, conduct slower, and exhibit higher heat
thresholds than CMH fibers (Weidner et al. 1999
). In
primates, they consist of type II A-fiber nociceptors that
are mechanically insensitive (Treede et al. 1998
).
More than 50% of the LT-THT neurons, however, started to respond when
the adapting temperature (35°C) changed by only 2°C (i.e., 37°C
in the heating direction and 33°C in the cooling direction). Since
nociceptors are unlikely to respond to such small changes in skin
temperature, we must propose that the initiation of the thermal
responses of LT-THT neurons is influenced by activation of cold and
warm receptors. A detailed examination of the responses during the
dynamic phase of the heat stimuli shows that the discharge rate of
LT-THT neurons increases in a near-linear fashion from 39 to 46°C,
and then decreases. This response profile is characteristic of C-warm
receptors (Duclaux and Kenshalo 1980; Hensel and
Iggo 1971
; Hensel and Kenshalo 1969
;
Konietzny and Hensel 1977
; Kumazawa and Perl
1977
; LaMotte and Campbell 1978
). Because some
LT-THT neurons started to respond when the temperature was lowered from 35 to 33°C, it is tempting to consider input from cold receptors as
well. A detailed examination of the responses during the dynamic phase
of the cold stimuli shows that the discharge rate of LT-THT neurons
decreases in a near-linear fashion from 20 to
10°C. This response
profile is characteristic of cold receptors (Dubner et al.
1975
; Duclaux et al. 1980
; Dykes
1975
; Hellon et al. 1975
; Hensel and Iggo
1971
; Iggo 1969
; Kenshalo and Duclaux
1977
; Poulos and Lende 1970a
,b
). For lack of
information about the response properties of these neurons to innocuous
cold stimuli (35-20°C), however, we cannot provide conclusive
evidence for this proposal.
Responses to stimulation of specific orofacial organs
Twenty-four percent of all THT neurons responded to stimulation of
the oral mucosa, tongue, or lips. They were located in the dorsomedial
region of laminae III, IV, and V of C1 and Vc; the principal termination area of primary afferent neurons that innervate the tongue, hard palate, and lips (Arvidsson et al. 1992, 1995
; Marfurt 1981
; Shigenaga et
al. 1986
). This distribution matches the distribution of
c-fos immunoreactive neurons following stimulation of the
tongue and lips (Carstens et al. 1995
; Strassman and Vos 1993
) and the recording sites of neurons responsive to noxious stimuli of other oral structures (Shigenaga et al.
1976
). In spite of the prominent presence of oral-sensitive
neurons in lamina I (Carstens et al. 1995
, 1998
;
Dostrovsky and Hellon 1978
; Hutchison et al.
1997
; Strassman and Vos 1993
), we did not find them. Three factors could contribute to our inability to record from
oral-sensitive neurons in lamina I; the use of relatively low-impedance
electrodes and their possible bias toward large neurons, the small
sample size (only 8 lamina I-THT neurons were recorded dorsomedially)
and the absence of lamina I-THT neurons that are oral sensitive.
Most (85%) oral-sensitive THT neurons encoded the intensity of the
noxious mechanical and thermal stimuli, over half (54%) encoded light
mechanical stimuli, and a few (15%) encoded innocuous cold. These
response properties of oral-sensitive THT neurons could be driven by
inputs they receive from nociceptors (Hayashi 1985;
Jacquin et al. 1986a
; Light et al. 1992
;
Sumino et al. 1973
), mechanoreceptors (Hensel and
Zotterman 1951
; Poulos and Lende 1970a
,b
), and
cold receptors (Dubner et al. 1975
; Heinz et al. 1990
; Hensel and Wurster 1970
; Hensel and
Zotterman 1951
; Poulos and Lende 1970a
,b
) that
innervate the tongue and the lips. Similar oral-sensitive nociceptive
(WDR and HT), mechanoreceptive (LT), and thermoresponsive Vc neurons
have been previously described (Carstens et al. 1998
;
Hoffman et al. 1981
; Hu et al. 1981
;
Hutchison et al. 1997
; McHaffie et al.
1994
; Price et al. 1976
; Renehan et al.
1986
), many of which project to the thalamus and brain stem. It
is therefore likely that oral-sensitive Vc neurons convey sensory
information to the hypothalamus which is similar to the information
they convey to other areas of the brain.
Sixteen percent of all THT neurons were classified as cornea-sensitive.
Since activation of the cornea by mechanical, thermal, or chemical
stimuli produces predominantly a sensation of pain in humans
(Beuerman and Tanelian 1979; Kenshalo
1960
; Lele and Weddell 1959
) and vocalization
and escape behaviors indicative of pain in animals (Gerard
1923
), cornea-sensitive neurons were considered nociceptive.
Given the many tasks of this study, however, it was not possible to
activate all classes of corneal receptors or to differentiate between
cornea alone and cornea plus conjunctiva stimulation for all but the
mechanical stimuli. Therefore the identification of cornea-sensitive
neurons was based only on their responses to cornea brush. The
application of a brush stimulus to the cornea is usually the most
effective way for identifying cornea-sensitive neurons as it activates
corneal A
mechanosensitive receptors (Belmonte et al.
1991
; Lele and Weddell 1959
; MacIver and
Tanelian 1993a
,b
; Tanelian and Beuerman 1984
)
and A
- and C-polymodal nociceptors (Belmonte and Giraldez
1981
; Belmonte et al. 1991
; Gallar et al.
1993
); receptors that are most easily excited by a moving
stimulus rather than by static corneal indentation (Belmonte and
Giraldez 1981
; Mosso and Kruger 1973
).
All cornea-sensitive THT neurons were recorded in ventrolateral
lamina V at the level of caudal Vc and rostral
C1, a gray matter area that receives input from
corneal nociceptors and periorbital receptors (Marfurt
1981; Marfurt and Del Toro 1987
; Panneton
and Burton 1981
; Shigenaga et al. 1986
) and
contains one of the two groups of cornea-sensitive TBNC neurons
(Bereiter et al. 1994
; Lu et al. 1993
;
Marfurt and Del Toro 1987
; Strassman et al.
1993
). While in this study all cornea-sensitive THT neurons
were found in lamina V, in previous studies they were found mainly in
lamina I (Carstens et al. 1998
; Hu et al.
1981
; Meng et al. 1997
; Nagano et al.
1975
; Nishida and Yokota 1986
; Pozo and
Cervero 1993
). Their absence from lamina I in this study was a
likely result of not recording there; a sample bias that often occurs
in electrophysiological experiments in which single-unit recording
techniques are used. The presence of cornea-sensitive neurons in lamina
V is not surprising, however, as it was demonstrated using
c-fos (Strassman et al. 1993
) and single-unit
recordings (Nagano et al. 1975
) in the rat.
About 19% of all THT neurons responded to electrical and mechanical
stimulation of the intracranial dura. Since pain is the only sensation
that can be evoked by stimulating the sinuses in the human, regardless
of whether the stimulus is electrical, mechanical, or chemical
(Ray and Wolff 1940), we have considered these neurons nociceptive. In fact, based on their responses to cutaneous
stimulation, most dura-sensitive THT units were classified as
nociceptive neurons capable of processing somatosensory signals that
originate in both the intracranial dura and extracranial (mainly
ophthalmic) skin. Their response properties suggest that the inputs
they receive originate in meningeal C and A
fibers that respond to
mechanical, thermal, and chemical stimulation (Bove and
Moskowitz 1997
; Strassman et al. 1996
), and
cutaneous nociceptors (discussed in the preceding text). The response
properties of these neurons resemble those of dura-sensitive neurons
that project to the thalamus in the cat (Davis and Dostrovsky
1988
). In fact, three of the dura-sensitive THT neurons also
project to the thalamus.
Responses of LRF-RHT neurons
All LRF-RHT neurons had large or complex receptive fields that extended beyond the innervation territory of the trigeminal nerve. They responded exclusively to noxious mechanical and thermal stimulation of extracephalic skin (e.g., paws and tail) and exclusively or preferentially to noxious mechanical and thermal stimulation of facial skin, lips and tongue and to brushing the cornea. The trends in their responses to mechanical and thermal stimulation of the facial skin (Fig. 13) suggest that they are capable of encoding the intensity of innocuous and noxious mechanical and noxious heat but not cold. Based on their response profiles, it is reasonable to propose that the inputs that drive them originate in nociceptive-specific (HT) spinal cord dorsal horn neurons and nociceptive (HT and WDR) plus non-nociceptive (LT) neurons in the TBNC.
In the absence of clear anatomical landmarks around the recording sites
that were found medial and ventral to the ventrolateral tip of nucleus
caudalis, we have opted to adapt Nord's nomenclature (Nord and
Kyler 1968) and define them as LRF neurons. This broadly defined area contains the caudal part of the A1 catecholaminergic cell
group (Dahlstrom and Fuxe 1964
), subnucleus reticularis
ventralis (Meessen and Olszewski 1949
), and possibly
a few subnucleus reticularis dorsalis neurons (Villanueva et al.
1988
). In agreement with our findings, this area was previously
found to receive direct input from nociceptive spinal cord neurons
(Craig 1995
; Lima et al. 1991
;
Menetrey et al. 1983
; Tavares et al.
1993
; Westlund and Craig 1996
), to contain
nociceptive neurons that respond to stimulation of orofacial organs
such as the cornea, skin, tongue, nose, and tooth-pulp in cats,
monkeys, and rats (Burton 1968
; Nagano et al.
1975
; Yokota et al. 1991
), and to project to the
hypothalamus (Cunningham and Sawchenko 1991
;
Loewy et al. 1981
; McKellar and Loewy
1981
; Sawchenko and Swanson 1981
) and forebrain
(Zagon et al. 1994
).
SUBNUCLEUS RETICULARIS VENTRALIS (SRV).
Although the recording locations of LRF-THT neurons resemble most
closely those of SRV neurons, we did not consider them to be typical
SRV units because of their receptive field size. The receptive fields
of SRV neurons are restricted to the innervation territory of the
trigeminal nerve (Burton 1968; Nagano et al. 1975
; Yokota et al. 1991
), while the receptive
fields of LRF-THT neurons often included the entire body. Many reasons
could account for this discrepancy. They include species differences,
exact recording locations, and the way in which studied neurons were selected (the only neurons we studied were those projecting to the hypothalamus).
SUBNUCLEUS RETICULARIS DORSALIS (SRD).
In spite of the close resemblance between the response properties of
the neurons we called LRF-RHT and those found in SRD (Villanueva
et al. 1988), we do not believe they were SRD units because
they were recorded caudal, ventral, and lateral to the SRD and because,
unlike SRD neurons, LRF neurons project to the hypothalamus.
CATECHOLAMINERGIC CELL GROUP (A1).
Anatomically, catecholaminergic neurons in the caudal ventrolateral
medulla are found as far dorsal as the recording locations of LRF-THT
neurons. Functionally, a comparison between A1 and LRF-RHT neurons
could not be completed at this point because little or no information
is available regarding the responsiveness of hypothalamic-projecting A1
neurons to innocuous and noxious somatosensory stimulation of organs
such as the skin, cornea, lips, and dura. Traditionally, A1 neurons
have been viewed as playing an important role in the reflex control of
hormonal secretion from hypothalamic neurons in response to
hemodynamic, gastrointestinal, and respiratory stimuli
(Cunningham and Sawchenko 1991; Randle et al.
1986
; Swanson 1987
; Willoughby et al.
1987
); sensory information they receive through the nucleus of
the solitary tract (Loewy 1990
). The finding that
hypothalamic projecting neurons in this region convey nociceptive information that originates in multiple organs indicates their potential involvement in the initiation of endocrine responses to
noxious stimuli as well; a consideration that was brought up recently
by Pan and colleagues using the Fos technique (Pan et al.
1999
).
Comparisons between THT and SHT neurons
Anatomically, the THT must be considered as the rostral extension
of the SHT. Together, the two tracts relay signals from all levels of
the spinal cord (Burstein et al. 1990) and therefore are
in a position to carry to the hypothalamus sensory information that
arises in the entire body. In the past 10 years, Giesler and colleagues
have studied the physiological properties of SHT neurons in the sacral
(Katter et al. 1996b
), lumbar (Burstein et al.
1991
), thoracic (Zhang et al. 1999a
), and lower
cervical (Dado et al. 1994b
) spinal cord. They found
that nociceptive information can be conveyed to the hypothalamus by
~90% of all SHT neurons (WDR and HT) and tactile information by
~50% of the neurons (WDR and LT). These neurons were capable of
processing sensory information from pelvic and abdominal organs such as
the vagina, colorectal canal, and bile duct, and from sacral, lumbar,
thoracic, and cervical dermatomes. It is therefore reasonable to
conclude that the spinohypothalamic and the trigeminohypothalamic
tracts are similar. Both SHT and THT neurons are located in the same
laminae, respond mainly to noxious stimuli but also to innocuous
stimuli, process somatosensory signals from cutaneous and visceral
organs, and exhibit receptive fields that, in general, are restricted
to no more than two to four dermatomes. The identification of
C1-2 and Vc neurons that project to the
hypothalamus and respond to noxious stimulation of intra- and
extracranial organs and trigeminal dermatomes is therefore not
surprising. A surprising finding regarding the physiological properties
of THT neurons is their ability to respond to mild mechanical and
thermal stimuli such as vibrissal deflections and skin cooling or
warming by <3°C. Three factors may explain the presence of these
responses in THT but not in SHT neurons: the unique innervation of
orofacial organs such as the tongue, lips, and nose by cold and warm
receptors and the vibrissae by specialized mechanoreceptors, the
application of most thermal stimuli to the hairy skin of the face in
this study versus the glabrous skin of the paws in the SHT studies, and
the use of slow ramps for thermal stimuli in this study versus fast
ramps in the SHT studies.
Based on the complete antidromic mapping of ascending and descending
axons of cervical SHT neurons (Dado et al. 1994a;
Kostarczyk et al. 1997
; Zhang et al.
1995
), partial mapping of lumbar (Burstein et al.
1991
) SHT axons, and mapping of a small number of sacral SHT
axons (Katter et al. 1996a
), it appears that most SHT
axons ascend to the hypothalamus through the contralateral lateral
funiculus and ventrolateral medulla. They then shift dorsally to create a tight bundle along the medial border of the medial geniculate nucleus. This tight bundle seems to loosen on entering the caudal diencephalon, where the axons shift ventrally to reach the supraoptic decussation through the zona incerta, cerebral peduncle, internal capsule, and optic tract. Like the SHT, THT axons ascend in the contralateral hypothalamus, cross the midline with the optic chiasm, and descend in the ipsilateral hypothalamus. As shown in Fig. 17, both
THT and RHT axons seem to follow SHT axons as they infiltrate through
the zona incerta, cerebral peduncle, internal capsule, and optic tract
and ascend in the SOD. Unlike SHT axons, however, THT and RHT axons
appear more scattered between the medial geniculate nucleus, the
central gray, and the substantia nigra. This difference suggests that
in the midbrain, ascending SHT axons may be partially segregated from
THT and RHT axons.
Functional considerations
In only 10 experiments, we successfully followed collateral
branches of parent axons from the supraoptic decussation to specific nuclei within the hypothalamus. Because anatomical studies have shown
that a large number of TBNC neurons project to the hypothalamus (Malick and Burstein 1998) and that many hypothalamic
nuclei contain trigeminal axons (Iwata et al. 1992
;
Newman et al. 1996
; Ring and Ganchrow
1983
), the scarcity of axonal branches in the hypothalamus suggests that the antidromic activation technique used in this study
was not adequate for detecting these branches. A likely explanation for
our inability to identify these axonal branches is that the stimulating
electrodes were too blunt (tip diameter, 35-50 µm) and the pulse
stimuli too short (200 µs) and too fast (10 Hz) to activate the small
(and probably unmyelinated) axonal branches. In fact, in the 10 experiments in which collateral branches were found in the
hypothalamus, sharper electrodes (tip diameter, 10-25 µm), wider
stimulus pulses (duration 200-800 µs), and slower interstimulus
intervals (1 Hz) were used. Collateral branches were found in the
anterior, lateral, and perifornical regions and the dorsomedial,
suprachiasmatic, and supraoptic nuclei.
Our findings raise the possibility that THT and RHT axons convey
trigeminal sensory signals to hypothalamic neurons that regulate body
temperature, food and water intake, sleep and circadian rhythms and a
wide range of behaviors (Bernardis and Bellinger 1993,
1998
; Kruk et al. 1983
; Lin et al.
1989
; Norgren 1970
; Panksepp
1971
; Peyron et al. 1998
; Roeling et al.
1993
; Saper 1995
; Scammell et al.
1993
; Sherin et al. 1996
; Simerly
1995
; Swanson 1987
). For example, many
low-threshold points of THT and RHT axons were identified within the
LH. Current understanding suggests that LH neurons play an important
role in the regulation of food and water intake, arousal, and
aggression (Bernardis and Bellinger 1993
; Date et
al. 1999
; Panksepp 1971
). Studies in which
lesions of the LH produced inhibition of food intake, and stimulation increased food intake, gave rise to the notion that LH neurons constitute a principle output pathway that promotes feeding behavior (Hetherington and Ranson 1940
; Stevenson
1970
). More recently, it has been proposed that LH neurons that
produce melanin-concentrating hormone or orexin (Bittencourt et
al. 1992
; Sakurai et al. 1998
) do so by
activating distinct autonomic, endocrine, and behavioral responses
through widespread projections to the cerebral cortex, brain stem, and
spinal cord (Elmquist et al. 1999
). Regarding water
intake, a recent study found that dehydration activates LH neurons and
produces a type of anorexia that can be reversed by sham drinking alone
(which does not affect water balance) (Watts 1999
). We
propose that THT neurons, which convey signals regarding the sensation
of water passing through the lips, oral cavity and esophagus, could
mediate the reversal of this dehydration-induced anorexia. The
locations of THT axons in the LH also correspond well with the location
of orexin-positive neurons (Date et al. 1999
). Because
of the putative role of orexin-positive neurons in the regulation of
sleep and arousal (Chemelli et al. 1999
; Lin et
al. 1999
; Peyron et al. 1998
), it is also
possible that THT neurons that relay nociceptive trigeminal signals can
contribute to changes in arousal states in response to pain.
Several THT axons also entered the SCN and SON. Because the SCN
regulates circadian rhythms such as sleep-wake cycles, plasma cortisol
levels, and body temperature (reviewed in van den Pol 1991), it is possible that the nociceptive signals that reach the SCN through the THT or RHT contribute to the disruption of normal
sleep patterns in pain patients. Because the SON produces oxytocin,
arginine-vasopressin, and corticotrophin-releasing hormone, which
regulate labor, lactation and stress responses (as reviewed in
Armstrong 1995
), it is possible that nociceptive signals
that reach the SON through the THT or RHT contribute to the initiation or cessation of these physiological functions by pain.
Trigeminohypothalamic tract neurons were also capable of transferring to the hypothalamus tactile information from oral and perioral organs such as the tongue, lips, and vibrissae. The physiological purposes for tactile signals reaching the hypothalamus are currently unknown and therefore a matter of speculation. They could be related to feeding, drinking, suckling, and a variety of intimate social behaviors such as kissing. For example, since rodents use the tongue, lips, and vibrissae to explore for food and water, sensory signals that provide information about the structure, size, consistency, and temperature of objects in the environment may play a role in the recognition of nutrients and the triggering (or avoidance) of feeding and drinking behaviors such as licking and chewing. In humans, it could be speculated that THT neurons capable of encoding cooling and warming sensations from the lips and tongue may be involved in the facilitation or suppression of the desire to eat foods that are colder or warmer than expected. Every-day life experience also points to the close association between the tactile sensation of dry lips and tongue and the desire to drink. Consideration can therefore be given to the idea that THT neurons may carry such sensations to hypothalamic areas that regulate osmotic and volemic states and by doing so, stimulate drinking behavior.
Forebrain projections
Because many spinal cord neurons project to forebrain nuclei
located anterior to the hypothalamus (Burstein and Giesler
1989; Burstein and Potrebic 1993
; Cliffer
et al. 1991
; Malick and Burstein 1998
), numerous
attempts were made by Giesler and colleagues to follow the axons of SHT
neurons from the SOD to the forebrain (Burstein et al.
1991
; Dado et al. 1994a
; Katter et al.
1996a
), but no such axons were found. This led to the
conclusion that SHT axons do not approach the forebrain through the
SOD. The present study confirms this conclusion. The few THT and RHT
axons that were followed in this study to the caudate-putamen, globus
pallidus, and substantia innominata ascended lateral to the
hypothalamus, within the internal capsule. A few anatomical studies
have documented the presence of axons in the forebrain that originate
in spinal (Cliffer et al. 1991
; Newman et al.
1996
), trigeminal (Newman et al. 1996
;
Yasui et al. 1987
), and LRF (Loewy et al.
1981
; Zagon et al. 1994
) neurons; findings
confirmed by injections of retrograde tracers in several forebrain
areas (Burstein and Giesler 1989
; Burstein and
Potrebic 1993
; Malick and Burstein 1998
;
Zagon et al. 1994
). Extracellular recording in CPu and
GP found two populations of neurons, those responding to innocuous
stimulation of orofacial receptive fields (Carelli and West
1991
; Levine et al. 1987
; Lidsky et al.
1979
; Schneider 1991
; Schneider et al.
1982
, 1985
) and those responding to noxious stimulation of the
entire body (Bernard et al. 1992
; Chudler et al.
1993
; Lin et al. 1985
; Richards and Taylor 1982
). Based on the involvement of CPu and GP neurons in the execution of motor functions, it was proposed that CPu and GP
neurons that respond to somatosensory stimulation may participate in
the coordination of complex motor responses to painful and tactile
stimuli (e.g., withdrawal and orientation). Regarding the ventral
pallidum/substantia innominata area, behavioral studies proposed that
these nuclei play a role in the generation of autonomic and somatomotor
aspects of emotional and motivational states (reviewed in Heimer
et al. 1997
). In 1992, Bernard et al. suggested that neurons in
this area receive nociceptive input and that this input arises in the
parabrachial nucleus. Our study suggests that nociceptive input to this
area can also arise in LRF-RHT neurons. Together, these nociceptive
pathways may contribute to the alteration of motivational state by pain.
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ACKNOWLEDGMENTS |
---|
We thank Dr. M. Jakubowski for valuable input and Dr. B. Ransil for the statistical analyses.
This work was supported by National Institutes of Health Grants DE-10904 and NS-35611-01, by the Education Fund of the Department of Anesthesia and Critical Care at Beth Israel Deaconess Medical Center, the Milton Fund, the Boston Foundation, the Goldfarb family, and the Fink family.
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FOOTNOTES |
---|
Address for reprint requests: R. Burstein, Dept. of Anesthesia and Critical Care, Harvard Institutes of Medicine, Room 830, 77 Avenue Louis Pasteur, Boston, MA 02115 (E-mail: rburstei{at}caregroup.harvard.edu).
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
Received 27 January 2000; accepted in final form 14 June 2000.
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REFERENCES |
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