Department of Physiology and Biophysics, Albert Einstein College of Medicine, Bronx, New York 10461
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ABSTRACT |
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Huang, Wu-Xin and Morton I. Cohen. Population and Unit Synchrony of Fast Rhythms in Expiratory Recurrent Laryngeal Discharges. J. Neurophysiol. 84: 1098-1102, 2000. In a decerebrate, vagotomized, gallamine-paralyzed cat that had a prominent bilaterally coherent fast rhythm (50 Hz) in expiratory (E) recurrent laryngeal (RL) nerve discharges, recordings were taken of the firing of nine RL E fibers. This rhythm (called E high-frequency oscillation or EHFO) was seen as a sharp peak in all unit autospectra, all unit-nerve coherence spectra (value range 0.39-0.91), and all unit-unit coherence spectra (value range 0.27-0.85). In addition, 8/9 units had a sharp autospectral peak in a lower frequency range (19-35 Hz) called E medium-frequency oscillation (EMFO), but there was no coherence at this frequency between signal pairs (unit-unit, unit-nerve, nerve-nerve). The MFOs are specific for each unit and are considered to arise from asynchronous inputs and membrane properties. The HFOs are considered to arise from widespread network interactions that produce a common (correlated) rhythm in virtually all neurons of the RL E network. These phenomena suggest the use of the RL E network as a model system for analyzing rhythmic neural interactions.
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INTRODUCTION |
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During activation of
a population of motoneurons, the spikes produced by different units are
synchronized on the time scale of the action, e.g., a limb movement or
a breath. But, in addition, different units' firing may be
synchronized on a faster time scale, as manifested by discharge rhythms
in the range 10-100 Hz (Farmer 1998).
In the respiratory system, an intrinsic fast rhythm (frequency range
50-100 Hz) is ubiquitous in inspiratory (I) motor nerve discharges
(phrenic, intercostal, laryngeal) and has been named high-frequency
oscillation (HFO) (Christakos et al. 1991, 1994
; Cohen et al. 1997
).
A similar type of bilaterally coherent oscillation (frequency range
24-54 Hz) was observed in recurrent laryngeal (RL) expiratory (E)
discharges and is called expiratory high-frequency oscillation (EHFO)
(Huang et al. 1993a). Interest in this type of rhythm
has been increased by our observation that similar EHFOs are evoked in
RL E discharges during fictive vocalization produced by midbrain (periaqueductal gray) stimulation (Nakazawa et al.
2000
).
The present study reports observations on EHFOs in discharges of
individual RL fibers that were recorded together with whole-nerve (population) RL discharges having this rhythm. Spectral analysis revealed that the rhythms of individual units were highly coherent with
(correlated to) the rhythms in other units' firing as well as with
rhythms in both ipsilateral and contralateral RL nerves. This finding
indicates that the EHFO originates from network interactions that
produce correlated outputs to RL motoneurons. A preliminary report of
this study was published in abstract form (Huang et al.
1993b).
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METHODS |
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Experimental preparation
The present study is based on data obtained in one cat of a
series of seven vagotomized cats (2.5-3.5 kg) that were decerebrated at the midcollicular level using standard methods (Kirsten and St. John 1978). Surgical preparation was performed under 3-5% halothane, the adequacy of anesthesia being attested by the absence of
movements and blood pressure changes. After decerebration and completion of surgery, the halothane was removed and the animals were
paralyzed by infusion of gallamine triethiodide (5 mg
kg
1
h
1). Recordings were not
started until at least 2 h after removal of halothane. A
pneumothorax was done and artificial ventilation was applied via a
mechanical ventilator connected to a 1-2 cm expiratory load. End-tidal
CO2 level (monitored by an infrared analyzer) was
maintained at 4-5% by varying ventilation or the composition of the
input gas mixture (0-5% CO2 in 100-95%
oxygen). To maintain fluid balance and keep systolic blood pressure
100 mmHg, the animals were perfused with 0.9% saline containing 5% glucose (4 mg kg
1
h
1). Rectal temperature
was kept at 37-39°C by use of a heating pad. At the end of the
experiment, the animals were given an overdose of sodium pentobarbital.
Recordings
Monophasic recordings were taken from one phrenic (PHR) nerve and from both the left and right RL nerves. (The RL nerve bundle on each side had been mobilized low in the neck after it separated from the main vagus trunk so that the vagi could be sectioned while leaving the RL nerve bundle centrally connected.) With the cat in a supine position, the severed nerves were mounted on bipolar electrodes and immersed in a mineral oil pool formed by a neck skin flap. Small bundles containing one or two active fibers were dissected from the right RL nerve and recorded with a monopolar electrode. As many as five isolated fibers from both I and E units were recorded simultaneously in different runs.
ANALOG SIGNALS. Whole-nerve and single-fiber (isolated bundle) discharges (preamplifier band-pass 1-5000 Hz), intratracheal pressure, and femoral arterial blood pressure were recorded on a videocassette recorder by means of a digital interface (Instrutech). The bandwidth was adequate for recording individual fiber spikes, which had a duration of ~1 msec.
PULSE SIGNALS.
Recordings were taken of pulses marking the onset of the I and E
phases, which were derived from PHR discharge by specialized circuitry
(Cohen 1968). Recordings were also taken of pulses
derived on-line from unit spikes by means of a time-amplitude
discriminator (Bak).
DIGITIZATION. Digitized recordings of the whole-nerve signals and of the pulses (as analog signals) were taken on-line via a personal computer-based analog-digital converter (RC Electronics). A sampling rate of 2500 Hz (0.4-msec bin duration) was used. Each of the 0.4-msec analog pulses was later converted by software to the number "1" in a data array. This time resolution was needed for the computation of autocorrelation histograms. Filtering before spectral computations was found to be unnecessary (as verified by test computations on filtered vs. unfiltered data) because the population (whole-nerve) signals had no periodic components in the 2500-5000 Hz range.
Data analysis
DATA WINDOWS.
Using the I and E pulse tag arrays derived from the phrenic potential
recording (Cohen 1968), portions of the data signal arrays corresponding to the I and E phases were identified. These were
used to mark data windows for correlation and spectral analysis. For
each recording, an ensemble of data windows was obtained that consisted
of segments corresponding to approximately the first half of the mean E
phase, which was the time during which E activity occurred in the RL
nerve (cf. Fig. 2A).
CYCLE-TRIGGERED HISTOGRAMS. Cycle-triggered histograms (CTHs) were computed with 40-msec bins, using the I or E pulses as triggers, for unit and rectified nerve activities. The latter were obtained by a digital algorithm that subjected the data to 40-Hz high-pass filtering followed by full-wave rectification.
AUTOCORRELATION HISTOGRAMS. Autocorrelation histograms (ACHs) were computed for unit pulses, using the original sampling bins of 0.4 ms, to verify that a recording had been obtained from a single unit, as was indicated by the absence of counts in the early portion (<2 ms) of the ACH, which corresponds to the refractory period.
FREQUENCY-DOMAIN ANALYSIS.
The fast Fourier transform was used to compute autospectra of the
signals and coherence spectra for pairs of signals (nerve-nerve, unit-unit, unit-nerve) using methods previously described
(Christakos et al. 1991). For the analog signals,
2.0-msec bins were used, the data value in each bin being derived by
summing the values in the original 0.4-msec sampling bins. This
procedure acted as a low-pass filter with an effective sampling rate of
500 Hz. The spike-derived pulse train for each unit was low-pass
filtered (cutoff, 250 Hz) by convolving the sequence of pulses with a
sinc function (defined as sin x/x) with appropriate parameters
(Christakos et al. 1984
).
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RESULTS |
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In the main series of seven decerebrate cats, only one had bilaterally coherent oscillations in RL E efferent nerve discharges; these EHFOs were present in six data runs recorded over a time period of 3.8 h. In these runs, recordings of activity in nine isolated RL E fibers were taken simultaneously with bilateral RL nerve recordings. During this series of recordings, there were no significant changes in the properties of the EHFOs in the nerve signals.
In six recording runs, one unit was recorded alone (single recording) and nine distinct unit pairs were recorded as follows: three were recorded together with one other unit (pair recordings) and six were recorded together with two other units (three triplet recordings in each of which the same unit pair was recorded together with an additional different unit).
For one of these triplet recordings the discharge patterns of the signals (bilateral RL E nerve activities, spike firing of 3 RL E units) during one respiratory cycle (I phase, time of phrenic firing; E phase, time of phrenic silence) are shown in Fig. 1A. It can be seen that during the first portion of the E phase there are prominent population waves (summed spike activities) in both RL nerves, as well as spike firing of the units.
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The temporal relations of population and unit activities are seen in the expanded traces of Fig. 1B, which shows that RL nerve activities are highly periodic (period of ~20 ms) and that the population waves are strongly synchronized between the two nerves, even though individual waves differ in amplitude. Of the 21 waves in the trace, 18 are closely synchronized to a spike of one or more units. Moreover, for 11/18 of these waves, the spikes of two units (8 cases) or three units (3 cases) are closely synchronized to each other (within 3 ms); these cases are labeled as coincident spike occurrence (black circles). For the remaining seven cases of unit-wave synchronization, only one unit's spike is synchronized to the wave (noncoincident spike occurrence, white circles).
Although every spike of each unit in Fig. 1B is synchronized to a population wave, the interspike interval durations have different 1:n relations to the population wave intervals. For example, interval 1 in the trace of unit 1 corresponds to three wave intervals (1:3 relation), interval 1 of unit 3 corresponds to two wave intervals (1:2 relation), and interval 2 of unit 3 corresponds to one wave interval (1:1 relation).
In Fig. 2A, the CTHs show the discharge patterns of phrenic and RL nerves and of units 1, 2, and 3. It can be seen that units 1 and 3 have a decrementing pattern whereas unit 2 has a bell-shaped pattern. The vertical brackets mark the duration of the windows (1.694 s, about half of the E phase duration) used in each cycle for gating the signals on which spectral computations were done.
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In Fig. 2B, the relations between unit 1 and nerve rhythms are indicated by the frequency components of the unit autospectrum (B1) and of the unit-(left RL) coherence spectrum (B2). In the unit autospectrum there are two prominent peaks at 22.9 and 48.8 Hz. However, the unit-nerve coherence shows no trace of the lower-frequency peak but has a prominent coherence peak (value 0.81) at the higher frequency, 48.8 Hz, as well as peaks at harmonics of that frequency. The higher-frequency component is therefore designated as EHFO whereas the lower-frequency component in the unit autospectrum resembles the medium-frequency oscillation (MFO) in fast inspiratory rhythms, which does not show coherence between different signals.
In all recordings, the left-right nerve coherence (LRL-RRL) at the dominant frequency (48-50 Hz) had a high peak value (>0.9) and did not show any lower-frequency (MFO) component. For all nine units the unit-nerve coherences (to both ipsi- and contralateral nerves) had high-coherence peak values (0.39-0.91) at the dominant frequency but did not have peaks at the lower (MFO-type) frequencies present in the unit autospectra.
The relations between rhythms of the three simultaneously recorded units of Fig. 1 are shown in Fig. 3. In the autospectrum for each unit (A1-A3) the first three peaks mark different frequency components. The first peak marks a component specific to each unit (22.8, 24.9, and 34.7 Hz for units 1-3, respectively). These components are not common to the different units, as indicated by their absence in the coherences between unit pairs (B1-B3) They may therefore be designated as MFO components. The second peak marks a frequency component (48-50 Hz) that is correlated between all unit pairs, as indicated by a prominent peak in each unit-unit coherence (B1-B3). Therefore this component is designated as HFO. The third peak marks a component that is the sum of the two preceding peaks and thus indicates an interaction between those two components. However, this combination frequency component is not coherent between unit pairs.
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All nine unit pairs had a prominent peak in the coherences at the HFO frequency (48-50 Hz). The range of peak values was 0.27-0.85. The MFO component, which was present in 8/9 units, had a frequency range of 19.0-35.2 Hz with mean of 26.4 ± 5.3 Hz (SD). Of these, six units had frequency peaks between 22 and 27 Hz. Finally, the combination frequency (peak 3, described in the preceding paragraph) was present in 7/9 units and had a frequency range of 70.3-85.0 Hz.
After these recordings there was a sudden change of state in which the coherent EHFOs disappeared in all RL E nerve and unit activities (this change was associated with a decrease of arterial blood pressure from 140/110 to 110/80 mmHg). During the new state, six runs were taken in which 10 RL E units were recorded. In this state, coherent EHFO peaks were absent in all RL nerve-nerve (left-right), unit-nerve, and unit-unit coherence spectra. However, the autospectrum for each unit's activity had a peak that was different for each unit (22-65 Hz) but that was not correlated to peaks in other autospectra. Among those units, one pair was recorded both before and after the change of state and showed this disappearance of EHFO. Thus comparison of unit discharge rhythms between the two states showed that the presence or absence of EHFOs did not arise from a selection bias in finding isolated units.
The HFOs present during the inspiratory phase (IHFO) in both phrenic
and RL I discharges had frequencies different from the EHFOs. In the
first 4 runs of the experiment, the IHFO frequency increased from 69 to
75 Hz and in the subsequent runs the frequencies remained stable in the
83-85 Hz range. This lack of relation between IHFO frequency and EHFO
frequency had been observed in other preparations that had EHFOs in RL
activity (Huang et al. 1993a).
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DISCUSSION |
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In this study we report that, in one preparation during the
time when strong fast rhythms were present in bilateral expiratory RL
nerve discharges, all individual RL E motoneurons (fibers) that were
recorded had synchronized discharges at a common frequency (48-50 Hz),
as indicated by the coherences between the activities of unit pairs and
between unit and nerve activities. We had previously described such
strong rhythmic synchronization between bilateral RL E nerve discharges
and designated the phenomenon as EHFO (Huang et al. 1993a).
Moreover, in the same preparation, when the EHFOs disappeared in the
nerve discharges they were also gone in unit discharges.
The strength of this fast rhythm in RL E nerve discharges (Fig.
1B) seemed greater than that of HFOs in inspiratory
(phrenic) activities reported in earlier studies (cf. Fig. 1 in
Cohen et al. 1997). The strength of this RL E population
rhythm seems to be based on its probable presence in the firing of
almost all individual RL E motoneurons, as suggested by the presence of
large autospectral peaks for all nine units of our sample taken during the time of occurrence of the population rhythm. The coherent frequency
component (48-50 Hz) may originate from the occurrence of interspike
intervals having values corresponding to the EHFO period (~20 ms,
reciprocal of 50 Hz) or the synchronization of individual spikes to the
periodic population bursts at the EHFO period, even if the interspike
intervals are not equal to the EHFO period.
In addition, 8/9 units had lower-frequency (MFO) autospectral peaks
(19-35 Hz; mean, 26 Hz) that were not coherent to frequency components
in this range in the autospectra of other units, as shown by the
absence of such peaks in the unit-unit and unit-nerve coherences. Such
components, which are now designated as EMFOs, arise in part from
interspike intervals that have values corresponding to the MFO period
(~40 ms, reciprocal of 25 Hz), as in the examples of Fig. 1. Since
each unit has a different MFO frequency, this rhythm seems to depend on
an individual unit's membrane properties as influenced by asynchronous
inputs. The independence of the MFO frequency of different units
results in the absence of unit-unit coherence in this frequency range.
Further, the summation of uncorrelated frequency components in the
population (nerve) recording results in cancellation and thus in the
absence of such components in the unit-nerve and nerve-nerve
coherences. It is of interest that an analogous phenomenon (correlated
IHFOs, noncorrelated IMFOs) is observed in inspiratory nerve activities
(Christakos et al. 1991; Cohen et al.
1997
).
It should be borne in mind that the linear spectral analysis done in this study would not necessarily detect nonlinear spectral relations (quadratic phase relations) between different frequency components. The presence of such relations is suggested by the existence of 1:n relations between unit and population (wave) intervals (Fig. 1B) and also by the existence of combination frequency peaks (HFO plus MFO) in unit autospectra (Fig. 3). Therefore the further elucidation of rhythmic relations will require the use of bispectral analysis.
The strength and prevalence of the EHFO coherences between different RL
E signals, and particularly between bilateral signals, indicates that
the oscillations arise from strong network interactions. The fact that
they are absent when RL E activity is smaller (Huang et al.
1993a), e.g., when vagal inhibition during I reduces I duration
and consequently E duration and RL E firing (Sica et al.
1985
), suggests that a critical level of excitation is needed for the activation of these feedback interactions. This idea is supported by the occurrence of prominent EHFOs in association with the
strong increase of RL E activity in fictive vocalization produced by
midbrain stimulation (Nakazawa et al. 2000
).
The prominence of the EHFOs in E RL activity indicates again the
ubiquity of this type of rhythmic interaction in neural networks. Because of the strength of the EHFOs during some states and the ease of
producing them by midbrain stimulation (Nakazawa et al. 2000), the RL E network may be a useful model system for
analyzing this type of fast oscillation.
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ACKNOWLEDGMENTS |
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This research was supported by National Heart, Lung, and Blood Institute Grant HL-27300.
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FOOTNOTES |
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Address for reprint requests: M. I. Cohen, Dept. of Physiology, Albert Einstein College of Medicine, 1300 Morris Park Ave., Bronx, NY 10461 (E-mail: mcohen{at}aecom.yu.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 1 February 2000; accepted in final form 24 April 2000.
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REFERENCES |
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