1Department of Anatomy and Neurosciences and 2Department of Internal Medicine, University of Texas Medical Branch, Galveston, Texas 77555
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ABSTRACT |
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Saab, Carl Y., Motohiro Kawasaki, Elie D. Al-Chaer, and William D. Willis. Cerebellar Cortical Stimulation Increases Spinal Visceral Nociceptive Responses. J. Neurophysiol. 85: 2359-2363, 2001. The role of the cerebellum in modulating nociceptive phenomena is unclear. In this study, we focus on the effects of cerebellar cortical stimulation on the responses of midline neurons of the lumbosacral spinal cord to graded nonnoxious and noxious visceral (colorectal distension) as well as somatic (brush, pressure, pinch) stimuli. Extracellular recording was used for the isolation and recording of spinal nociceptive neurons, while electrical current pulses and chemical injection of D, L-homocysteic acid were used to stimulate the cortex of the posterior cerebellar vermis. Cerebellar cortical stimulation increased the responses of all isolated cells to colorectal distension, whereas the effect on the responses to somatic stimuli was variable. These findings indicate that the posterior cerebellar vermis may exert a pro-nociceptive effect on spinal visceroceptive neurons.
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INTRODUCTION |
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The cerebellum is endowed with a
highly regular circuitry often regarded as a micro-processing system
(Arbib 1987; Eccles et al. 1967
;
Middleton and Strick 1998
). This system plays a
recognized role in motor phenomena (Dow and Moruzzi
1958
; Fields and Willis 1970
; Gilman et
al. 1981
; Holmes 1922
, 1939
; Palay and
Chan-Palay 1982
) but a controversial role in pain
(Spiegel 1982
; Wu and Chen 1990
). To
clarify the role of the cerebellum in nociception, it is necessary to
verify that the cerebellum receives an input triggered by noxious
events and that a cerebellar output modulates nociceptive phenomena.
Nociceptive inputs to the cerebellum have been reported previously.
Electrophysiological data indicate that surface potentials recorded
from the cerebellar cortex are evoked following stimulation of C fibers
in the saphenous (Wu and Chen 1990) and radial nerves (Ekerot et al. 1991
). Electrical stimulation of the
cerebellar vermis causes antidromic activation of neurons in the
lateral medullary reticular nucleus that respond to noxious colorectal distention (CRD) (Ness et al. 1998
). Axons containing
substance P, a peptide neuromodulator of nociceptive transmission, were found to form contacts with neurokinin-1 receptor-immunoreactive cells
in the dorsal spinocerebellar tract that project to paravermal areas
(McGonigle et al. 1996
). Imaging of the human brain in
conscious individuals reveals blood volume increases in the cerebellar
vermis or regions more lateral to it during acute heat pain
(Casey et al. 1994
), the warm-discrimination task
(Casey et al. 1996
), muscle pain (Svensson et al.
1997
), and capsaicin-evoked pain and allodynia (Iadorola
et al. 1998
). Similarly, an increase in activity was observed
in the cerebellar vermis following intradermal capsaicin injection in
the rat hindpaw using electrophysiologic and functional magnetic
resonance imaging techniques (Saab et al. 1999
).
Cerebellar stimulation affects pain-related behavior. Anti-nociceptive
effects were reported following electrical stimulation of the brachium
conjunctivum in the monkey (Siegel and Wepsic 1974);
similar observations were made after microinjection of morphine into
the anterior cerebellum of the rat (Dey and Ray 1982
).
Interestingly, it has been postulated that impulses generated by
posterior column stimulation may lead to relief of pain and spasticity
by activating the cerebellum and thus affecting "not only [its]
static innervation, but also the central conduction and perception of
pain impulses" (Spiegel 1982
). Nevertheless cerebellar
modulation of spinal responses to noxious stimuli has not, to our
knowledge, been investigated yet.
In this study, we have examined the influence of cerebellar cortical stimulation on spinal nociceptive neurons that responded to noxious visceral and somatic stimuli. Our results indicate that stimulation of the cerebellar vermis may lead to a pro-nociceptive effect at the level of the spinal cord.
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METHODS |
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Thirteen adult male Sprague-Dawley rats were used in this study.
All experimental procedures were approved by the Institutional Animal
Care and Use Committee and were in accordance with the guidelines of
the National Institutes of Health and the International Association for
the Study of Pain. The rats were anesthetized with pentobarbital sodium
(60 mg/kg ip). Supplementary infusion of pentobarbital sodium (5 mg · kg1 · h
1) was maintained through catheterization of
the jugular vein. Respiration was assisted by a tracheal cannula, and
body temperature was kept at 37°C using a heating blanket. A
laminectomy and a craniotomy exposed the lumbosacral spinal cord and
the posterior cerebellum, respectively. The rat was then fixed on a
stereotaxic apparatus (Kopf Instruments), and a balloon was gently
inserted through the rectum for visceral stimulation by CRD. The dura
over the cerebellum and the spinal cord was carefully excised to permit stimulation and recording, respectively.
Visceral stimulation
CRD is a well-documented visceral stimulus (Al-Chaer et
al. 1996a,b
, 1998
; Su and Gebhart 1998
). The
balloon was constructed from a latex glove finger attached to a
4-cm-long tygon tubing that was connected to a manual pump and a
pressure transducer to monitor stimulus intensity. CRDs consisted of
consecutive inflations to pressures ranging from 20 to 80 mmHg applied
in increments of 20 mmHg for 20 s every 4 min. CRDs of 60 or 80 mmHg using this balloon are considered noxious (Al-Chaer et al.
2000
).
Somatic stimulation
The cutaneous stimuli that were applied consisted of brushing
(BR) the receptive field (RF) using a camel hair brush, applying pressure (PR) to a fold of skin using a large arterial clip, or pinching (PI) with a small arterial clip exerting a force of
approximately 550 g/mm2. All these stimuli are
considered as nonnoxious except PI, which is distinctly painful if
applied to a fold of human skin (Dougherty et al. 1992).
Twenty seconds of background activity was recorded preceding each
stimulation. BR, PR, and PI were applied consecutively for 10 s
separated by 10 s of no stimulation.
Extracellular recording
Extracellular single-unit recordings were made stereotaxically
from the lumbosacral spinal cord (1-1.7 mm from the surface, near the
midline) using a tungsten microelectrode (0.5-µm diameter tip,
12-M impedance). The extracellularly recorded action potentials were
evoked by either CRD or somatic stimuli applied to the skin, then
amplified, fed into a window discriminator, and displayed on an
oscilloscope. The outputs from the window discriminator and from the
amplifier were led into a data-collection system (CED 1401+) and a
computer for data compilation as "rate histograms" using the Spike
2 software program. The responses to CRDs were calculated as the
differences between the mean rate of firing during a stimulus and that
during the 20 s baseline recording that preceded each CRD. The
responses to cutaneous stimuli were calculated similarly as the
differences between the mean rate of firing during a stimulus and that
during the 10 s that preceded each stimulus.
Cerebellar stimulation
A tungsten microelectrode was guided vertically by a micromanipulator into the posterior cerebellar vermis (lobule VI) until the first layer of spontaneously active cells was encountered (1-2 mm from the vermal surface). The electrode was then disconnected from the recording equipment and connected instead to a Master 8 stimulator for the generation of current pulse trains (0.2-ms pulses at 333 Hz, 100-150 µA). Electrical stimulation started at 2 s before the start of a stimulus and lasted for either 10 s during each CRD or 5 s during each somatic stimulation. Therefore electrical stimulation was terminated at 10 or 5 s before the end of a CRD or a somatic stimulation, respectively. This paradigm was advantageous in allowing for observation of the effect of electrical stimulation alone on the baseline (2 s prior to CRD), and arguing against an electrical stimulus artifact (during CRD).
For chemical stimulation, a 10-µl Hamilton microsyringe was filled with a 0.1 M solution of D,L-homocysteic acid (DLH) and advanced stereotaxically into the same region as described above for electrical stimulation. Ten seconds before each CRD, 2 µl DLH was injected, whereas only one 3-µl injection was applied 10 s before the start of each somatic stimulation session.
Histology
Small lesions were made by passing DC (15-20 µA for 5 s) through the microelectrode to mark the recording site in the spinal cord (Fig. 1A) and the stimulation site in the cerebellum (Fig. 1B). The tissue was then preserved in 10% neutral formalin, then prepared for sectioning (50-µm thick) using a microtome. Sections were stained using neutral red.
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RESULTS |
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Fifteen units were isolated. All units were activated by CRD, while seven were also activated by applying cutaneous stimuli on the hindlimbs or the pelvic region. The isolated neurons had a low level of background discharges (0-10 spikes/s) and were classified as "nociceptive" based primarily on their responses to CRD at intensities higher than 40 mmHg but also on their responses to noxious cutaneous stimuli where applicable. Accordingly, all cells were classified as nociceptive even though some also responded to nonnoxious stimulation.
Neuronal response characteristics
Cerebellar stimulation caused significant increases in the responses of cells to all intensities of CRD (Figs. 2B and 3). Facilitation occurred in nine neurons using only chemical injections compared with six using only electrical current. However, when the percent increase in the responses to graded visceral stimuli was compared between the two neuronal populations (chemical versus electrical stimulation) using a t-test, the two groups were not found to be significantly different (P > 0.5). Therefore data from both groups were pooled in Fig. 3. However, variable effects were observed on the responses to somatic stimuli (Table 1). For example, the responses to PR and PI applied on the hindlimb of one rat were increased (Table 1, rat I and Fig. 2C), but only the responses to BR were increased in another rat while those to PR and PI were decreased (Table 1, rat II). Applied alone, cerebellar stimulation did not seem to have an effect on the background discharges of cells (Fig. 2C4). In addition, some responses to graded CRDs and somatic stimuli were tested twice before cerebellar stimulation to rule out any sensitization effect that may have been caused by repetitive noxious stimulation. Repeated noxious stimulation did not alter the response pattern (examples of repetitive visceral and somatic stimuli are shown in Fig. 2, A and C, 1 and 2, respectively). In one rat, responses to graded CRDs applied alone 20 min after cerebellar stimulation with DLH were still significantly increased compared with the initial responses, indicating that the cerebellar effects may be long lasting (data not shown).
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DISCUSSION |
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The brain regions involved in the processing of nociceptive
information are often described as sensory regions because pain is
first and foremost a sensory experience, and in parallel, a motor
function has been attributed to the cerebellum. However, many
objections have been raised to the conceptual schism between sensory
and motor phenomena (Fetz 1992), especially in the
cerebellum (Bloedel and Bracha 1995
; Houk
1997
; Schmahmann 1991
, 1997
; Schmahmann and Pandya 1997
), which seems to play a role in both cognitive (Fiez 1996
; Schmahmann and Sherman 1997
)
and nociceptive information processing (Wu and Chen
1991
).
In this study, we have shown that stimulation of the cerebellar cortex
increases the responses of spinal nociceptive neurons to noxious, as
well as nonnoxious, CRDs and often modulates their responses to
different somatic stimuli. The disparity in the effects of cerebellar
stimulation on visceral versus somatic spinal inputs may be due to the
disparity in the effects of cerebellar stimulation on visceral versus
somatic spinal pathways (for example, Sluka et al. reported in 1997 that intradermal injection of capsaicin in monkeys caused a significant
increase in the responses of spinothalamic cells to weak mechanical
stimuli but not to noxious mechanical or heat stimuli). Furthermore it
is not clear whether the increase in the responses to nonnoxious
stimuli observed in this study implies that the same stimuli may
provoke a nociceptive reaction based solely on these data. If so, then
this reaction should produce allodynia in the unanesthetized state,
whereas increases in the responses to noxious stimuli should result in
hyperalgesia. The neurons were isolated from the midline zone of the
spinal cord near the central canal. This region contains cells of
origin of a newly identified component of the postsynaptic dorsal
column (PSDC) pathway that largely conveys visceral input to the dorsal column nuclei (Al-Chaer et al. 1996a,b
). In fact, the
PSDC was suggested to contribute, at least in part, to the somatic
nociceptive input to the cerebellum (Ekerot et al.
1991
). However, it is not clear whether both visceral and
somatic information is relayed via the same pathway to the cerebellum.
When the effects of cerebellar stimulation are appraised at the spinal
or peripheral levels, they are described as "rather complex"
(Eccles et al. 1967, p. 257). This complexity is due to
the fact that even focal electrical stimulation excites not only
Purkinje cells, the sole output from the cerebellar cortex, but also
granule cells and their axons, basket, stellate, and Golgi cells and
climbing and mossy fibers, as well as mossy and climbing fiber
collaterals that may activate the inferior olive and many subcortical
nuclei. In this study, the chemical injection of DLH may have limited
the activation to cortical cells around the injection site by only
depolarizing cell bodies of Purkinje cells and cerebellar interneurons
but not axons coursing en passant (Goodchild et al.
1982
), such as climbing or mossy fibers. The effects of
electrical and chemical stimulation in this study were not
significantly different.
The enhancement of nociceptive responses observed in this study is
consistent with the analgesic effects previously described following
cerebellar cortical lesions (Chambers and Sprague
1955a,b
; Russel et al. 1894
; but also refer to
Bloedel and Bracha 1995
). Moreover, an interesting
observation made by Siegel and Wepsic (1974)
points to
an alteration of nociceptive thresholds by electrical stimulation of
the cerebellum in monkeys. These authors found that a profound and
long-lasting analgesia was produced on activation of the brachium
conjunctivum. However, stimulation of lobulus simplex (lobule VI) and
other regions of the posterior cerebellum resulted in decreased
thresholds for withdrawal from a noxious stimulus. In another study in
which the effect of cerebellar lesions on the potency of
morphine-induced analgesia was tested in rats, it was reported that
lesions of the anterior cerebellum markedly decreased the duration of
analgesia caused by systemic administration of morphine (Dey and
Ray 1982
). However, prolongation of the analgesic effects
following posterior cerebellar lesions was also noted. In light of
these and our findings, it is likely that the anterior cerebellum may
be exerting an overall anti-nociceptive effect. By contrast, the
posterior cerebellum decreases the latency of withdrawal from noxious
stimuli and enhances spinal nociceptive responses.
Cerebellar modulation of spinal cord nociceptive responses and
pain-behavior must depend on descending pathways to the spinal cord
(Mendlin et al. 1996; Palay and Chan-Palay
1982
; Voogd and Glickstein 1998
). Anatomical
interconnections exist between the cerebellum and the reticular
formation of the brain stem, nucleus raphé magnus, locus
c
ruleus, and the periacqueductal gray region, all considered to be
involved in descending inhibition or "sensory gating." In this
case, stimulation of the cerebellar cortex may inhibit the deep
cerebellar nuclei projecting to these brain stem centers, thus
relieving the spinal nociceptive neurons from tonic inhibition. If
cellular responses to CRD and somatic noxious stimuli can be used as
indices for pain, then there seems to be a cerebellar role in the
modulation of pain.
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ACKNOWLEDGMENTS |
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The authors thank Dr. Y.-C. Park Arai for assistance in the later stages of this study.
This study was supported by National Institute of Neurological Disorders and Stroke Grants NS-11255 and NS-09743.
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FOOTNOTES |
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Address for reprint requests: W. D. Willis, University of Texas Medical Branch, 301 University Blvd., Galveston, TX 77555-1069 (E-mail: wdwillis{at}utmb.edu).
Received 4 December 2000; accepted in final form 14 March 2001.
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REFERENCES |
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