Department of Pharmacology and Toxicology, Michigan State University, East Lansing, Michigan 48824-1317
![]() |
ABSTRACT |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Gebber, Gerard L., Sheng Zhong, Craig Lewis, and Susan M. Barman. Differential Patterns of Spinal Sympathetic Outflow Involving a 10-Hz Rhythm. J. Neurophysiol. 82: 841-854, 1999. Time and frequency domain analyses were used to examine the changes in the relationships between the discharges of the inferior cardiac (CN) and vertebral (VN) postganglionic sympathetic nerves produced by electrical activation of the midbrain periaqueductal gray (PAG) in urethan-anesthetized, baroreceptor-denervated cats. CN-VN coherence and phase angle in the 10-Hz band served as measures of the coupling of the central oscillators controlling these nerves. The 10-Hz rhythm in CN and VN discharges was entrained 1:1 to electrical stimuli applied to the PAG at frequencies between 7 and 12 Hz. CN 10-Hz discharges were increased, and VN 10-Hz discharges were decreased when the frequency of PAG stimulation was equal to or above that of the free-running rhythm. In contrast, stimulation of the same PAG sites at lower frequencies increased, albeit disproportionately, the 10-Hz discharges of both nerves. In either case, PAG stimulation significantly increased the phase angle between the two signals (VN 10-Hz activity lagged CN activity); coherence values relating their discharges were little affected. However, the increase in phase angle was significantly more pronounced when the 10-Hz discharges of the two nerves were reciprocally affected. Importantly, partialization of the phase spectrum using the PAG stimuli did not reverse the change in CN-VN phase angle. This observation suggests that the increase in the CN-VN phase angle reflected changes in the phase relations between coupled oscillators in the brain stem rather than the difference in conduction times to the two nerves from the site of PAG stimulation. In contrast to the effects elicited by PAG stimulation, stimulation of the medullary lateral tegmental field induced uniform increases in the 10-Hz discharges of the two nerves and no change in the CN-VN phase angle. Our results demonstrate that changes in the phase relations among coupled brain stem 10-Hz oscillators are accompanied by differential patterns of spinal sympathetic outflow. The reciprocal changes in CN and VN discharges produced by PAG stimulation are consistent with the pattern of spinal sympathetic outflow expected during the defense reaction.
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
In addition to exerting uniform actions on the
discharges of sympathetic nerves, the brain can formulate differential
patterns of spinal sympathetic outflow. For example, the defense
reaction is characterized, in part, by increased sympathetic drive to
the heart, vasoconstriction in the viscera, and skeletal muscle
vasodilation due to the withdrawal of sympathetic vasoconstrictor
discharge and activation of sympathetic vasodilator fibers
(Coote et al. 1973; Folkow et al. 1964
;
Hilton 1982
). The underlying mechanisms responsible for
increased sympathetic drive to the heart and viscera in the face of
decreased vasoconstrictor outflow to skeletal muscle remain obscure.
Two models have attracted the most attention in recent years. The first
model is based on the observation that chemical activation of the
rostral ventrolateral medulla elicits site-dependent differential
changes in regional blood flows (Dampney and McAllen
1988
; McAllen and Dampney 1990
). This has led to
the proposal that presympathetic neurons in the rostral ventrolateral medulla are topographically organized according to their peripheral targets. A logical extension of this proposal is that the discharges of
such neuronal groups are coordinated by inputs from the defense regions
of the midbrain periaqueductal gray (PAG) and hypothalamus (Carrive 1991
; Dampney 1994
). In the
second model, the discharges of selected groups of preganglionic
sympathetic neurons with different targets are coordinated by inputs
from brain stem and hypothalamic neurons with diffusely branching
spinal axons. Jansen et al. (1995)
have proposed that
such inputs, some of which arise in the defense regions of the PAG and
hypothalamus, act as "command" neurons that, when activated, lead
to differential patterns of spinal sympathetic outflow.
The models described above differ in terms of the level of the neuraxis at which cell groups with different targets are coordinated. Nevertheless, both models presuppose coordination as the result of inputs shared by groups of neurons that are not necessarily directly interconnected. The implication is that such inputs excite some cell groups and inhibit others. The current study deals with an alternative to these "hard-wired" models. In the alternative model, differential patterns of spinal sympathetic outflow elicited in the absence of baroreceptor reflex feedback control are emergent properties of a system of dynamically coupled brain stem oscillators that generate the 10-Hz rhythm in sympathetic nerve discharge (SND).
The background for this model is as follows. First, a 10-Hz rhythm
uncorrelated to rhythms of similar frequency in the
electroencephalogram (EEG) and motor systems (inferior olivary
activity) is ubiquitous to the discharges of sympathetic nerves with
cardiovascular targets in decerebrate or urethan-anesthetized cats
(Barman et al. 1992, 1995
; Gebber
et al. 1994b
). The 10-Hz rhythm in SND is most prominent after
baroreceptor denervation or unloading (Barman and Gebber 1997
; Barman et al. 1992
, 1994
).
Second, the 10-Hz rhythm is generated in the brain stem by multiple
oscillators, each of which preferentially or selectively controls a
different portion of the spinal sympathetic outflow (Gebber et
al. 1994b
; Huang et al. 1992
). Third, whereas the 10-Hz rhythmic discharges of sympathetic nerves with different targets normally are strongly correlated, the coupling of the brain
stem oscillators controlling the nerves is both dynamic and nonuniform
(Barman et al. 1992
; Gebber et al.
1994b
).
In the current study, we tested whether electrical activation of selected sites in the brain stem of baroreceptor-denervated cats reciprocally affects the 10-Hz discharges of the inferior cardiac (CN) and vertebral (VN) postganglionic sympathetic nerves, and if so, whether such changes are accompanied by alterations in coherence and phase angle in the 10-Hz band. Coherence and phase angle were used as measures of the relationships among the coupled oscillators controlling these nerves. The CN and VN innervate the heart and forelimb vasculature, respectively. The results demonstrate that differential patterns of spinal sympathetic outflow emerge when the phase relations among coupled 10-Hz oscillators are altered by stimulation of the defense region of the midbrain PAG.
![]() |
METHODS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Experimental subjects and anesthesia
The protocols used on 15 cats were approved by the
All-University Committee on Animal Use and Care of Michigan State
University. The cats were initially anesthetized with 2.5% isoflurane
mixed with 100% oxygen. Urethan (1.2-1.8 g/kg iv) was then
administered, and isoflurane inhalation was terminated. This dose range
of urethan has been reported to maintain a surgical level of anesthesia
for a period (8-10 h), which exceeded the duration of our experiments (Flecknell 1987). Nevertheless, supplemental doses (0.2 g/kg iv) of urethan were given every 4-6 h. The frontal-parietal EEG
showed a mixture of 7- to 13-Hz spindles and delta-slow waves,
indicative of unconsciousness and blockade of information transfer
through the thalamus (Steriade and Llinas 1988
). The EEG
was not changed by noxious stimuli (e.g., pinch) applied to the head or
body and, as previously reported, was not correlated to 10-Hz or lower
frequency SND (Barman et al. 1995
).
General procedures
Blood pressure was measured from a catheter inserted into the
abdominal aorta via a femoral artery. Spontaneous respiration during
anesthesia was eupneic with end-tidal CO2
(Traverse Medical Monitors Capnometer, model 2200) in the normocapnic
range. Subsequently, the animal was paralyzed (gallamine triethiodide;
4 mg/kg iv, initial dose), artificial ventilation with room air was
begun, and bilateral pneumothoracotomy was performed. End-tidal
CO2 was kept between 4.0 and 4.5% by adjusting
the parameters of artificial ventilation. Rectal temperature was kept
near 38°C with a heat lamp. Baroreceptor denervation was performed by
bilateral section of the carotid sinus, aortic depressor, and vagus
nerves (Barman et al. 1992). Section of these nerves
eliminated the cardiac-related rhythm in SND and the inhibition of SND
induced by raising blood pressure with a bolus iv injection of
norepinephrine bitartrate (2 µg/kg).
Neural recordings and central stimulation
By using methods described by Barman et al.
(1992), potentials were recorded monophasically with bipolar
platinum electrodes from the central ends of the cut CN and VN near
their exits from the left stellate ganglion. The CN and VN provide
sympathetic outflows to the heart and forelimb vasculature,
respectively. Nerve recordings initially were made with the band-pass
of the Grass Instruments 7P3 preamplifier set at 1-1,000 Hz, so that envelopes of multiunit spikes appeared as slow waves (Barman et al. 1992
; Cohen and Gootman 1970
). The
frontal-parietal EEG was recorded with a gold-plated disk electrode
placed on the skull and the indifferent electrode on crushed muscle;
the amplifier band-pass was 1-1,000 Hz. These data were stored on
magnetic tape.
A Grass S8800 quartz-timed digital stimulator and PSIU6
constant-current unit were used to deliver 1-ms square-wave pulses of
variable intensity and frequency through concentric bipolar stainless
steel electrodes (Rhodes model SNE-100 with 0.25-mm tip exposures
separated by 0.75 mm) to selected sites in two regions of the brain
stem. Electrode placements into the midbrain PAG and medullary lateral
tegmental field (LTF) were made visually after removal of portions of
the occipital and parietal bones and medial cerebellum. Relative to the
stereotaxic coordinates of Snider and Niemer (1961),
electrode placements in the midbrain PAG were caudal to the bony
tentorium at P1 to P2, L1 to L2 (left), and H + 3 to H + 0. We refer to
this portion of the midbrain PAG as the caudal PAG. The sites of
stimulation in the medullary LTF were located 2-4 mm rostral to the
obex, 2-3 mm lateral to the midline (left), and 2-4 mm below the
dorsal surface. Sites of stimulation were identified with references to
electrode tracks. The brain stem was removed and fixed in 10% buffered
Formalin. Frontal sections of 30-µm thickness were cut on a cryostat,
stained with cresyl violet and examined microscopically.
Data analysis
A flow chart depicting the methods and sequence of data analysis
is presented in Fig. 1. The recordings of
SND on magnetic tape were initially low-pass filtered at 50 Hz to
obtain the "original" records shown in Fig. 1A. The
analog filter had an attenuation slope of 24 dB/octave. In preparation
for time series analysis, the original records were then digitally
filtered without phase distortion to extract the 10-Hz band of SND.
This was performed by using software obtained from R. C. Electronics, Santa Barbara, CA. The width of the band-pass for digital
filtering usually was 4 Hz with the center frequency matched to that of
the primary peak in the autospectrum of SND. The digitally filtered
signals (Fig. 1B) are smoother and more sinusoidal-like than
the originals with power reduced by no more than 10% in the designated
band-pass. The roll-off slope of the digital filter was such that power
outside of the band-pass setting was reduced by 39%/Hz. By using
software written by one of us (Lewis), voltages and times of occurrence of the peaks and troughs of the digitally smoothed signals were detected and saved as an ASCII file. These data were used to construct time series of the amplitudes (peak to trough) of the slow waves in the
two nerves, and the relative phase (i.e., phase angle in degrees)
between the peaks of the CN and VN slow waves on a cycle-by-cycle basis
(Fig. 1C). Slow-wave amplitudes were normalized (scale
0-1.0) relative to the largest slow wave in the time series. The phase angle between the peaks of corresponding CN and VN slow waves was
plotted on a scale of 0-360° with the peak of the CN slow wave in
each cycle serving as the reference. Thus the phase angle is the number
of degrees that the peak of the VN slow wave lagged that of the CN slow
wave. Phase angle () was calculated on a cycle-by-cycle basis by
using the formula
![]() |
|
Spectral analysis was used to characterize CN and VN discharges
(original recordings) in the frequency domain. This provided information not available in the time series such as the values of
power in different frequency bands and coherence values relating signal
pairs. Fast Fourier transform (FFT) was performed by using a modified
version (Gebber et al. 1994b) of the programs of
Cohen et al. (1987)
and Kocsis et al.
(1990)
. The sampling rate was 200 Hz, and the resolution of
measurement was 0.2 Hz/bin. Thirty-two 5-s windows were averaged with
50% overlap for 80-s data blocks and 75% overlap for 40-s data
blocks. The analysis yielded autospectra of the discharges of the CN
and VN (Fig. 1D, top 2 panels) and corresponding coherence
functions (Fig. 1D, 3rd panel) and phase spectra (Fig.
1D, bottom) relating the discharges of the two nerves. In
each panel, the spectra for data collected before (control; trace
1 or
for phase angle) and during (test; trace 2 or
) brain stem stimulation are superposed. The autospectrum of a
signal shows how much power (voltage squared) is present at each
frequency. Powers in the superposed autospectra are normalized on a
scale of 0-1.0, with 1.0 representing the highest absolute power
reading found in one of the bins of either the control or test
spectrum. The coherence function (normalized cross-spectrum) measures
the strength of linear correlation (scale 0-1.0) of pairs of signals as a function of frequency, whereas the phase spectrum measures the lag
(scale 0-360°) of the second signal (VN) in a pair relative to the
first (CN) at each frequency. Although FFT was performed for the 0- to
100-Hz band, the spectra are displayed on a frequency scale of 0-15
Hz. Less than 10% of the total power in SND was contained above 15 Hz.
In this study, the total power in SND is defined as the sum of the
absolute values in the bins between 0 and 15 Hz. A macro written in
Microsoft Excel version 7.0 was used to measure the power above
background in the 10-Hz band. A line was fitted to connect the left and
right limits of the sharp peak near 10 Hz in the autospectrum of SND.
The power in the 10-Hz band was calculated as the area above the line.
The power at frequencies between 0 and 6 Hz is the sum of the values in
the bins comprising this band. Changes in power are expressed as a
percent of control. Because the range over which power could change was
not restricted to values between 0 and 100%, paired and unpaired
comparisons were made with the Student's t-test using raw
percentages (Sokal and Rohlf 1969). However, coherence
values were z-transformed (Rohlf and Sokal
1969
) before paired comparisons were made. Values in the text
are means ± SE with P
0.05 used to signify
statistically significant differences. The Pearson product-moment
correlation coefficient (r value) (Rohlf and Sokal
1969
) was used to test for a significant relationship between
the changes in sympathetic nerve powers and CN-VN lag time produced by
brain stem stimulation.
Partialization of the CN-VN coherence function and phase spectrum was
performed by using standardized 5-V square-wave pulses (5-ms duration)
representing electrical stimuli applied to the PAG. This procedure
allowed us to determine whether the responses of the two nerves (locked
1:1 to the stimuli) were attributable to factors in addition to the
direct influences of their inputs from the PAG. As described in earlier
reports from our laboratory (Gebber et al. 1994a,b
),
partialization involves, first, mathematical removal of the portion of
two signals (CN and VN discharges) that is attributable to a third
signal (PAG stimulus), and second, computation of the relationships
between the residual components of CN and VN discharges. The
mathematical formulas used to calculate the partial coherence and
partial phase spectrum, and the algorithms on which our software is
based can be found in Jenkins and Watts (1968)
and
Bendat and Piersol (1986)
. If the coherence of CN and VN
discharges is attributable solely to the direct influences of inputs
from the PAG, partialization will reduce the CN-VN coherence value at
the frequency of stimulation to zero. If, on the other hand, the
central circuits governing these nerves are tightly coupled,
partialization will not necessarily reduce the coherence value to zero
(Gebber et al. 1994a
,b
; Kalitzin et al.
1997
; Lopes da Silva et al. 1980
). In such
cases, we also partialized the CN-VN phase spectrum using the PAG
stimuli. This allowed us to test whether the changes in CN-VN phase
angle (10-Hz band) produced by PAG activation reflected alterations in
the phase relations among coupled oscillators rather than the
difference in conduction times to the two nerves from the site of
stimulation. In the former case, the phase angle during PAG stimulation
should be the same before and after partialization (Jenkins and
Watts 1968
). In the latter case, the change in phase angle
produced by PAG stimulation would be reversed by partialization if the
residual coherence value in the 10-Hz band remained significantly
different from zero. The value of phase angle would be random and,
thus, meaningless if coherence is reduced to zero.
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Effects of caudal PAG stimulation on 10-Hz rhythm in SND of baroreceptor-denervated cats
SINGLE SHOCKS.
The biphasic primary response elicited in both the CN and VN by single
shocks (100-500 µA) applied once every 2 s to sites in the
caudal PAG was an increase in activity (upward negative potential)
followed by a decrease in activity (downward positive wave). A typical
example is shown in Fig. 2, where the
traces are peristimulus averages of 122 CN and VN responses with the PAG stimulus applied at time 0. In 9 cases, the onset
latency of the negative potential was 64 ± 4 (SE) ms for the CN
and 74 ± 6 ms for the VN. The ratio of the negative to positive
wave (peak amplitudes measured from baseline) was 1.7 ± 0.2 for
the CN and 0.9 ± 0.1 for the VN. The durations of the positive
waves in the CN (200 ± 25 ms) and VN (192 ± 21 ms) were not
significantly different. As shown in Fig. 2, the biphasic primary
responses of the CN and VN were followed by damped oscillations whose
periods corresponded to that of the free-running (no stimulation) 10-Hz rhythm. The damped oscillations indicate that PAG stimulation reset the
10-Hz rhythm in SND (Huang et al. 1992; Pavlidis
1973
). The portions of the averages of CN and VN activities
preceding the PAG stimuli are flat because the stimuli were delivered
randomly with respect to the phases of the free-running 10-Hz rhythm.
|
FREQUENCIES OF STIMULATION (7-12 HZ) NEAR THAT OF THE FREE-RUNNING RHYTHM. The responses of the CN and VN were related to the frequency of PAG stimulation in a remarkable way. Frequencies of PAG stimulation at or slightly above that of the free-running rhythm produced reciprocal changes in the 10-Hz discharges of the two nerves, whereas lower frequencies of stimulation of the same sites did not. In the experiment illustrated in Fig. 3, the frequency of caudal PAG stimulation was the same as that (10.0 Hz) of the free-running rhythm in SND. The amplitude-time series show that PAG stimulation (begun at the 1st vertical line) almost immediately increased 10-Hz slow-wave amplitude in the CN (Fig. 3A, top) but decreased that in the VN (Fig. 3A, middle). The changes in slow-wave amplitudes were accompanied by an increase in the phase lag of VN 10-Hz activity relative to CN 10-Hz activity from between 40 and 90° to between 110 and 180°. The change in phase angle is shown in both the relative phase-time series (Fig. 3A, bottom) and the short strips of digitally filtered records of CN and VN discharges (Fig. 3, B and C) that formed part of the data block from which the time series were derived. Note that the effects of PAG activation were reversed soon after PAG stimulation was stopped at the second vertical line in Fig. 3A.
|
|
|
![]() |
|
|
|
|
|
|
HIGH-FREQUENCY STIMULATION. High-frequency (25 Hz) PAG stimulation reciprocally affected the 10-Hz discharges of the CN and VN. A typical example is shown in Fig. 10. As demonstrated with time series (Fig. 10A) and spectral (Fig. 10B) analyses, CN 10-Hz discharges were increased and VN 10-Hz discharges were decreased during high-frequency stimulation. Moreover, CN-VN phase angle in the 10-Hz band was increased. Note that the frequency of the free-running rhythm was not changed by high-frequency PAG stimulation (Fig. 10B, top 2 panels), and there was little change in the coherence values in the 10-Hz band (Fig. 10B, 3rd panel). CN-VN phase angle at the frequency of peak coherence in the 10-Hz band was increased from near 80° to near 160° (Fig. 10B, bottom panel).
|
Stimulation of the medullary LTF
The effects on CN and VN discharges produced by electrical activation of eight sites in the medullary LTF (see METHODS) of six baroreceptor-denervated cats were strikingly different from those produced by PAG stimulation. Independent of whether the LTF stimulus frequency (7-14 Hz range) was below, equal to, or higher than that of the free-running rhythm, CN and VN 10-Hz band powers were increased to nearly the same extent (Fig. 11, A-C, top 2 panels). Moreover, phase angle in the 10-Hz band was essentially unchanged (Fig. 11, A-C, bottom panel). Peak coherence in this band also was little affected by LTF stimulation (Fig. 11, A-C, 3rd panel). As previously described for the PAG, peak power in the 10-Hz band was moved to frequencies of LTF stimulation that were below (Fig. 11A) or above (Fig. 11C) that of the free-running rhythm. Thus LTF as well as PAG stimulation entrained the 10-Hz rhythm in SND. Although not shown, high-frequency (25 Hz) LTF stimulation increased CN and VN discharges uniformly and raised mean blood pressure by >40 mmHg.
|
Our summary of the effects of medullary LTF stimulation is limited to eight episodes when the stimulus frequency was the same as that of the free-running rhythm (e.g., Fig. 11B). Stimulus intensity ranged from 150 to 500 µA. Power in the 10-Hz band of CN activity was increased to 363 ± 39% of control, whereas VN 10-Hz power was increased to 392 ± 66% of control. CN-VN phase angle at the frequency of peak coherence in the 10-Hz band was not significantly affected. Values of phase angle were 40 ± 23° before and 54 ± 26° during medullary LTF stimulation. Peak coherence values in the 10-Hz band were 0.93 ± 0.02 before and 0.96 ± 0.01 during medullary LTF stimulation.
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Reciprocal changes in sympathetic outflows to the heart (or
kidney) and skeletal muscle in response to midbrain PAG or hypothalamic stimulation were first reported by Kollai and Koizumi
(1980) and Dean and Coote (1986)
. These studies,
however, were performed without reference to the frequency composition
of SND or the phase relations between the discharges of nerve pairs.
Moreover, no consideration was given to the question of whether the
pattern of changes produced by central activation was dependent on the frequency of stimulation. The current study focused on these issues.
As did Kollai and Koizumi (1980) and Dean and
Coote (1986)
, we used electrical stimuli to activate sites in
the brain. It is obvious that electrical stimulation is an artificial
means of eliciting sympathetic nerve responses. Nevertheless, this
approach allowed us to uncover the dependency of pattern on stimulus
frequency, thus providing a window through which underlying mechanisms
for the reciprocal changes in CN and VN activities could be examined. Although we cannot attribute the changes in SND produced by electrical stimulation specifically to the activation of neuronal somata, enhanced
cardiac function and reciprocal changes in blood flows to the viscera
and skeletal muscle have been reported on chemical activation of
neurons in the caudal PAG (Carrive 1991
; Dampney 1994
; Hilton and Redfern 1986
).
Previous work from our laboratory demonstrated that the 10-Hz rhythm in
SND is generated in the brain stem by a system of coupled oscillators,
each of which preferentially or selectively controls a different
portion of the spinal sympathetic outflow (Gebber et al.
1994b; Huang et al. 1992
). The current study
extends these findings in new directions. First, we found that this
system of coupled oscillators is influenced by inputs from the defense region of the caudal PAG. Regarding this point, single shocks applied
to the PAG reset the 10-Hz rhythm in SND. Furthermore, peak power in
the 10-Hz band of SND was moved to the frequency of PAG stimulation
(range, 7-12 Hz) at the expense of power at the frequency of the
free-running rhythm. We interpret this finding to indicate that inputs
from the PAG entrained the 10-Hz rhythm (Glass and Mackey
1988
; Huang et al. 1992
; Pavlidis
1973
). Second, we found that the differential changes in CN and
VN discharges elicited by PAG stimulation were largely restricted to
the 10-Hz band of activity. Regarding this point, PAG stimulation
either failed to significantly affect the power in the 0- to 6-Hz band of CN and VN discharges or, in one experimental series, changed 0- to
6-Hz power in CN discharges in a direction opposite that of the change
in 10-Hz band power. Thus the differential changes in CN and VN
discharges were attributable primarily to perturbation of the system
responsible for the 10-Hz rhythm.
Concerning the mechanisms responsible for the reciprocal changes in CN
and VN 10-Hz discharges, several observations argue against the
possibility that PAG stimulation simply engaged hard-wired pathways
that independently excited some brain stem oscillators or spinal cell
groups while inhibiting others. First, single shocks applied to the
caudal PAG elicited short-latency excitatory responses in both the CN
and VN. Second, whereas VN 10-Hz activity was significantly reduced by
PAG stimulation at frequencies equal to or above that of the
free-running rhythm, stimulus frequencies just below that of the
free-running rhythm significantly increased the 10-Hz discharges of
this nerve. The critical stimulus frequency at which the reversal occurred always was that of the free-running rhythm. This observation is remarkable considering the fact that the frequency of the
free-running rhythm ranged from 8.0 to 11.8 Hz. Third, the coherence
value relating CN and VN discharges at the frequency of PAG activation (7-12 Hz range) remained significantly different from zero in most
cases following partialization using the PAG stimuli. The residual
coherence indicates that the 10-Hz oscillators controlling the two
nerves remained tightly coupled during their entrainment in a 1:1
relation to the PAG stimuli (Gebber et al. 1994a,b
;
Kalitzin et al. 1997
; Lopes da Silva et al.
1980
). Importantly, the phase angle relating CN and VN
discharges at the frequency of PAG stimulation was not changed after
partialization. Thus lengthening of the CN-VN phase angle during PAG
stimulation could not be attributed simply to the difference in
conduction times to the two nerves from the site of stimulation.
Rather, lengthening of the phase angle suggests that PAG stimulation
altered the phase relations between the coupled 10-Hz oscillators
governing the CN and VN. This suggestion is further supported by the
results obtained with high-frequency PAG stimulation. As did
frequencies of stimulation at or slightly above that of the
free-running rhythm, high-frequency (25 Hz) stimulation reciprocally
affected CN and VN discharges and lengthened the phase lag of VN 10-Hz
activity relative to CN activity. In the case of high-frequency
activation, it is unlikely that lengthening of the CN-VN phase angle
can be explained by differences in conduction times to the two nerves
from the site of stimulation because the 10-Hz rhythm was not entrained
1:1 to the PAG stimuli.
It is also unlikely that PAG stimulus-induced decreases in VN 10-Hz
activity reflected postexcitatory depression that was more pronounced
in the VN than CN. If postexcitatory depression was an important factor
in explaining the reduction of VN 10-Hz activity, we would not have
expected the frequency of PAG stimulation at which the reversal from
increased to decreased VN 10-Hz activity occurred to be strictly tied
to the frequency of the free-running rhythm. Moreover, we would not
have expected LTF stimulation to increase CN and VN 10-Hz discharges
proportionately if postexcitatory depression was more pronounced in the
VN than CN. Finally, cycle-by-cycle measurements of slow-wave amplitude
demonstrated that the inhibition of VN 10-Hz discharges produced by PAG
stimulation was not preceded by an initial excitation (see Figs.
1C and 3A). Although Kollai and Koizumi
(1980) reported that the duration of postexcitatory depression
following hypothalamic stimulation was longer in the VN than CN of the
cat, we did not find this to be the case when single shocks were
applied to the PAG once every 2 s. Nevertheless, we did find that
the ratio of amplitudes of the initial negative wave and succeeding
positive wave elicited by single shocks was greater for the CN than VN.
Coherence and phase angle were used to characterize the coupling of the 10-Hz discharges of the CN and VN. The former measures the strength of coupling, whereas the latter measures the relative timing of the bursts in the two nerves. Whereas the strength of coupling was not much affected by PAG stimulation, the reciprocal changes in CN and VN 10-Hz discharges were accompanied by a dramatic increase in the phase lag of VN activity relative to CN activity. Under the assumption that the state of coupling of brain stem circuits is reflected by the CN-VN phase angle, the results of the current study are consistent with the hypothesis that dynamic changes in the phase relations between coupled 10-Hz oscillators lead to differential patterns of spinal sympathetic outflow. First, the differential changes in the 10-Hz discharges of the two nerves produced by PAG stimulation were temporally correlated to the increase in the phase lag of VN activity relative to CN activity. Second, the magnitude of change in phase angle was directly related to the extent to which PAG stimulation differentially affected the 10-Hz discharges of the two nerves. Thus the phase angle was increased more when CN and VN 10-Hz discharges were reciprocally affected than when the 10-Hz discharges of both nerves were increased, albeit disproportionately. Third, uniform increases in the 10-Hz discharges of the CN and VN produced by medullary LTF stimulation, and occasionally by frequencies of PAG stimulation below that of the free-running rhythm, were not accompanied by a significant change in phase angle in the 10-Hz band.
The question can be raised whether the changes in CN-VN phase angle
produced by PAG stimulation reflected changes in the shape of the 10-Hz
sympathetic nerve slow waves (e.g., from sinusoidal to asymmetric
spikelike waves) rather than alterations in the phase relations between
coupled brain stem oscillators. This possibility seems unlikely for two
reasons. First, time series analysis was performed only after digital
filtering was used to extract the 10-Hz slow waves from the total
signal. The filtered slow waves were sinusoidal in shape both before
and during PAG stimulation (see Figs. 1B and 3, B
and C). Second, in the case of spectral analysis, FFT
determines the phase angle by fitting series of harmonically related
sine and cosine waves to the data (Bendat and Piersol
1986; Jenkins and Watts 1968
).
Whereas our hypothesis is that changes in the phase relations among
coupled 10-Hz oscillators reflect the events leading to differential
patterns of spinal sympathetic outflow, it remains to be determined
whether phase angle itself is the parameter responsible for the
patterns that emerge. Discussion of this matter could be useful in
guiding the direction of future studies. In general terms, coupled
nonlinear oscillators are dynamical systems in that changes in the
levels of control parameters (inputs to system) lead to abrupt changes
in the relative timing (i.e., phase relations) of their outputs during
each cycle of activity (Haken 1996; Kelso 1995
). The phase relations characterizing the state of coupling in which the system exists at any given time is determined by the
internal self-organizing properties of the system and the values of the
control parameters. The repertoire of possible states is dependent, in
addition, on such factors as the number of oscillators in the system
and their interconnections (e.g., near vs. far neighbors). These
factors remain to be defined for the brain stem system responsible for
the 10-Hz rhythm in SND. Models of such systems have been used to
explain how the pattern of locomotion might be changed from one
characterized by in-phase movements of the limbs to another characterized by out-of-phase movements (Beek et al.
1992
; Grillner 1981
; Haken 1996
;
Jeka et al. 1993
; Kelso 1995
). Here, we
invoke the same general principles to explain how a shift in phase
angle might lead to reciprocal changes in sympathetic outflows to
different targets.
Whereas skeletal muscle contractions generally follow the
frequency of rhythmic motor nerve activity, this is not the case for
vascular smooth muscle. Rather, vascular smooth muscle acts as a
low-pass filter with a cutoff frequency well below that of the 10-Hz
rhythm. How then might changes in the phase relations among coupled
10-Hz oscillators in the brain stem lead to differential patterns of
spinal sympathetic outflow and, thus, changes in regional blood flow
occurring in opposite directions? Because elimination of the 10-Hz
rhythm in SND by chemical inactivation or ablation of selected brain
stem regions significantly reduces blood pressure (Zhong et al.
1993), it seems reasonable to assume that 10-Hz SND is
transduced into a "steady-state" level of vascular smooth muscle
contraction. Under this condition, differential patterns of spinal
sympathetic outflow would result if alterations in the phase relations
among coupled oscillators induced nonuniform changes in the amplitudes
of the 10-Hz slow waves in nerves with different targets. Regarding
this possibility, in-phase bidirectional coupling of two oscillators by
excitatory connections can lead to mutual reinforcement of their
outputs during each cycle of rhythmic activity (König and
Schillen 1991
). This situation can be changed by incorporating a lag into the system. For example, Reddy et al. (1998)
have demonstrated that coupled limit cycle oscillators can drive one
another to a state of zero amplitude output when their mutual
interactions are suitably delayed. Perhaps most relevant to the current
study is a model of oscillators coupled by mutual inhibitory
connections (Glass and Mackey 1988
). One of the possible
functional states in such a system is characterized by a high rate of
firing of the cells of one of the oscillators and a low rate of firing
of cells in the second oscillator. The applicability of such models to
the data reported in the current study remains to be determined.
Chemical or electrical activation of the caudal PAG elicits changes in
heart rate and regional blood flows characteristic of the naturally
occurring defense reaction (Carrive 1993; Dampney 1994
; Hilton 1982
; Hilton and Redfern
1986
). In cats, full expression of the increase in blood flow
to skeletal muscle during the defense reaction requires not only the
activation of sympathetic cholinergic vasodilator fibers and the
release of epinephrine from the adrenal medulla, but also the selective
inhibition of vasoconstrictor outflow to these vascular beds
(Coote et al. 1973
; Folkow et al. 1964
).
Thus the reciprocal changes in power in the 10-Hz band of CN and VN
discharges, and the change in CN-VN phase angle produced by caudal PAG
stimulation may represent, in part, the electrophysiological correlate
of the defense reaction. This suggestion is complicated by the fact
that the VN innervates the vasculature of forelimb skin as well as
skeletal muscle (Kollai and Koizumi 1980
). Clearly, additional experiments are needed to test whether the 10-Hz rhythm in
SND is involved in mediating the cardiovascular components of the
defense reaction.
The small but statistically significant increase in blood pressure accompanying the reciprocal changes in CN and VN 10-Hz discharges produced by PAG stimulation was not unexpected. If the changes for these nerves were generally representative of those for sympathetic outflows to the viscera and skeletal muscle, then increased nerve activity directed to the viscera would be balanced, in part, by decreased activity to skeletal muscle. However, the rise in blood pressure produced by frequencies of PAG stimulation that significantly increased the 10-Hz discharges of both nerves was similar to that observed during the reciprocal changes elicited by slightly higher frequencies of stimulation of the same sites. This apparent discrepancy can be explained if one assumes that the total power in SND is a better predictor of blood pressure than 10-Hz band power. Regarding this point, the changes in total power of CN discharges were similar in the two experimental groups. Moreover, the changes in CN and VN total power were small relative to the changes in the 10-Hz band. This is because power in the 0- to 6-Hz band of SND was little affected by PAG stimulation.
In summary, our results support the view that the reciprocal changes in the 10-Hz discharges of the CN and VN produced by caudal PAG stimulation in baroreceptor-denervated cats are emergent properties of a system of coupled brain stem oscillators with different targets. Specifically, we propose that alterations in the phase relations among coupled 10-Hz oscillators reflect the events leading to differential patterns of spinal sympathetic outflow. The proposal we have raised represents a considerable departure from classical views that are based on the premise that the formulation of differential patterns of spinal sympathetic outflow simply involves the activation of hard-wired pathways that independently excite certain cell groups in the brain stem and/or spinal cord while inhibiting others. Rather, we view inputs to the coupled 10-Hz oscillators as functioning to lead the system through a repertoire of internally self-organized states, each of which is characterized by a different set of phase relations and, thus, a different pattern of spinal sympathetic outflow.
![]() |
ACKNOWLEDGMENTS |
---|
The authors thank S. Sykes for typing the manuscript.
This study was supported by National Heart, Lung, and Blood Institute Grant HL-13187.
![]() |
FOOTNOTES |
---|
Address reprint requests to G. L. Gebber.
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 7 January 1999; accepted in final form March 1999.
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|