Two distinct phases characterize maturation of neurons in the nucleus of the tractus solitarius during early development: Morphological and electrophysiological evidence

We have used electrophysiology and light microscopy of intracellularly labeled neurons in the nucleus of the tractus solitarius (nTS) in brainstem slices of the newborn rat (PO to P6) to examine the functional and morphological correlation of their development. Three-dimensional reconstruction of neurons injected intracellularly with biocytin, following electrophysiological recording, revealed a close correspondence between morphological immaturity (appearing as polarization of the dendritic tree) and the absence of a ramp-like voltage trajectory at the offset of hyperpolarizing current injections–l_A negativity (8 of the 8 cells examined showed this correlation). These morphologically polarized I_A negative neurons showed preferential dendritic sprouting in two diametrically opposite poles of the perikaryon. The orientation of the polarity differed according to the rostrocaudal location of the neuron. The appearance of a polarized dendritic tree during the first (immature) phase was transient and closely coincident with I_A negativity. Following the development of adult-like electrophysiological characteristics, i.e., I_A positivity, nucleus of the tractus solitarius neurons showed remarkably different morphological features (9 of 10 cells). These included a widespread branching of the dendritic tree in all directions, giving it a bushy appearance (cell body to dendrite ratio of 1:40). Numerous dendritic spines, growth cones on both dendrites and axons, and axon collateralization were present during both phases and indicate that nTS neurons during the two phases of early development demonstrate dynamic features of growth and maturation. The development of adult-like electrophysiological characteristics, i.e., I_A positivity, progressively increased in the postnatal period. During the later part of the first postnatal week, twice as many neurons showed I_A positivity in days P3 to P6 as compared with days PO to P2. These results reveal the dynamic nature of neurons in the nTS during early development and illustrate the close link between morphology and functional characteristics in this region. We suggest that the establishment of adult-like morphology can be modified by appropriate environmental clues provided to nTS neurons during the initial (immature) phase of early postnatal development. © 1993 Wiley-Liss, inc.     

Rhythmic neuronal activities in the nucleus of the tractus solitarius isolated in vitro

Coronal slices of the rat medulla were used for an electrophysiological study of the nucleus of the tractus solitarius isolated in vitro. Half of the spontaneously active neurons in the ventral NTS exhibited a rhythmic repetitive spike discharge. In contrast, bursting activity was virtually absent in the isolated NTS. Resetting of repetitive discharges could be obtained by electrical stimulation of the glossopharyngeal-vagal afferent terminals in the tractus solitarius. These results are discussed in relation to the proposed role of the NTS in the neurogenesis of the respiratory rhythm.     

Substance P innervation of neurons projecting to the paraventricular hypothalamic nucleus in the rat nucleus tractus solitarius

After injection of WGA-HRP-colloidal gold in the rat paraventricular nucleus (PVN), retrogradely labeled neurons were found mainly in the medial and commissural subnuclei of the nucleus tractus solitarius (NTS) around 0.5 mm caudal to the obex which is closely related to cardiovascular function. Electron microscopic immunohistochemistry in these areas demonstrated synaptic contacts between retrogradely labeled neurons and substance P-immunoreactive terminals. Innervation of NTS-PVN projection systems by substance P is suggested.     

Rapidly adapting pulmonary receptor afferents: I. Arborization in the nucleus of the tractus solitarius

The organization of axon collaterals, preterminal processes, and presumptive synaptic boutons of single physiologically identified rapidly adapting receptor (RAR) pulmonary afferent fibers was examined following the intraaxonal application of wheat germ agglutinin conjugated with horseradish peroxidase (WGA-HRP). The RAR axons were injected 200-300 microns lateral to the nucleus of the tractus solitarius (nTS) at a number of different rostrocaudal levels in seven individual experiments. The trajectories of the stained axons were reconstructed from individual 50-microns-thick serial sections. The rostrocaudal extent, as well as the distribution of the trajectory of each RAR afferent, was reconstructed from every section by using a camera lucida attachment. In this first of two papers, we describe the pattern of organization of bouton terminals of RAR afferents related to cytoarchitectonically distinct subnuclei of the nTS. In the companion paper, morphological details of the fine structure of these synaptic boutons and axonal branches are described in different subnuclei in order to illustrate morphological differences in these functionally distinct regions. A number of significant findings have resulted from this light microscopic study. The central process of a single RAR afferent fiber arborized in the medulla oblongata over a considerable distance in the rostrocaudal plane (2.5 mm rostral to 1.4 mm caudal to the obex). A single RAR afferent fiber terminated in numerous bouton terminals (range 500-1,050), and these terminals arose from over 400 segments of branches of the parent injected axon. A small number of en passant bouton terminals were found. There appeared to be a remarkable degree of consistency in the subnuclei of the nTS where these terminals arborized. The dorsal and dorsolateral subnuclei of the nTS received 144-647 bouton terminals. The second-largest concentration of bouton terminals of RAR afferents was found in the intermediate (nI) subnucleus of the nTS. No labeled bouton terminal was found in the ventral and ventrolateral subnuclei of the nTS. This finding is in sharp contrast to the terminations of SAR afferents which terminated predominantly in the ventral and ventrolateral nuclei of the nTS, the interstitial nucleus of the nTS, and the nI. The parent RAR axon could be traced as far rostrally as 2.5 mm, even though the region of terminal arborization could not be followed beyond 0.8 mm. The destination of this rostrally projecting RAR afferent could not be determined in this study.(ABSTRACT TRUNCATED AT 400 WORDS)     

Direct synaptic projections to esophageal motoneurons in the nucleus ambiguus from the nucleus of the solitary tract of the rat

Neurons of the nucleus of the solitary tract (NTS) serve as interneurons in swallowing. We investigated the synaptology of the terminals of these neurons and whether they project directly to the esophageal motoneurons in the compact formation of the nucleus ambiguus (AmC). Following wheat germ agglutinin conjugated horseradish peroxidase (WGA-HRP) injection into the NTS, many anterogradely labeled axodendritic terminals were found in the neuropil of the AmC. The majority of labeled axodendritic terminals (89%) contained round vesicles and made asymmetric synaptic contacts (Gray's type I), but a few (11%) contained pleomorphic vesicles and made symmetric synaptic contacts (Gray's type II). More than half of the labeled terminals contacted intermediate dendrites (1-2 μm diameter). There were no retrogradely labeled medium-sized motoneurons, but there were many retrogradely labeled small neurons having anterogradely labeled axosomatic terminals. A combined retrograde and anterograde transport technique was developed to verify the direct projection from the NTS to the esophageal motoneurons. After the esophageal motoneurons were retrogradely labeled by cholera toxin subunit B conjugated HRP, the injection of WGA-HRP into the NTS permitted ultrastructural recognition of anterogradely labeled axosomatic terminals contacting directly labeled esophageal motoneurons. Serial sections showed that less than 20% of the axosomatic terminals were labeled in the esophageal motoneurons. They were mostly Gray's type I, but a few were Gray's type II. In the small neurons, more than 30% of axosomatic terminals were labeled, which were exclusively Gray's type I. These results indicate that NTS neurons project directly not only to the esophageal motoneurons, but also to the small neurons which have bidirectional connections with the NTS. J. Comp. Neurol. 381:18-30, 1997. © 1997 Wiley-Liss, Inc.     

Ultrastructural characterization of pharyngeal and esophageal motoneurons in the nucleus ambiguus of the rat

The viscerotopic organization of the upper alimentary tract has been established in the nucleus ambiguus, but there is little information about the morphology of the individual neurons innervating the pharynx and esophagus. We studied the ultrastructure of pharyngeal (PH), cervical esophageal (CE), and subdiaphragmatic esophageal (SDE) motoneurons labeled by retrogradely transported wheat germ agglutinin conjugated horseradish peroxidase (WGA-HRP) in the compact formation of the nucleus ambiguus. WGA-HRP was injected into the lower pharynx, or the cervical and subdiaphragmatic esophagus of male rats. The retrogradely labeled PH neurons in the rostral portion of the compact formation were large (26.1 × 50.1 μm, 906.7 μm²), polygonal, and contained well-developed cell organelles with a round nucleus. Subsurface cisterns connected with rough endoplastic reticulum were often present near the postsynaptic membrane. Both CE and SDE neurons in the compact formation were medium-sized, round or oval, and contained well-developed cell organelles, although the SDE neuron was significantly larger than the CE neuron (24.9 × 33.6 μm, 593.0 μm² in the SDE neuron, and 19.5 × 30.2 μm, 440.3 μm² in the CE neuron). The average number of axosomatic terminals in a sectional plane was largest in PH neurons (29.0), smaller in CE neurons (7.9), and smallest in SDE neurons (4.2). The number of axosomatic terminals containing round vesicles (Gray's type I) was almost equal to that of terminals containing pleomorphic vesicles (Gray's type II) in PH and CE neurons, but there were few Gray's type II axosomatic terminals in SDE neurons. Desmosome-like junctions at somato-somatic or somato-dendritic apposition were often present in the area surrounding SDE neurons. There were also small unlabeled neurons (9.5 × 18.1 μm, 131.8 μm²) in the compact formation of the nucleus ambiguus. The small neurons contained poorly developed cell organelles and an irregular shaped nucleus with invaginated nuclear membrane, and had no Nissl bodies. These results indicate that PH neurons have the characteristics of somatic motoneurons, and that CE and SDE neurons are similar to visceral motoneurons. © 1996 Wiley-Liss, Inc.     

Morphology of inspiratory neurons located in the ventrolateral nucleus of the tractus solitarius of the cat

The morphology of 11 dorsal respiratory group (DRG) inspiratory neurons located in the ventrolateral nucleus of the solitary tract (vl-NTS) was studied using the technique of intracellular labeling with the enzyme horseradish peroxidase (HRP). Six of these cells were cut in the transverse plane and had a mean somal diameter of 30.4 m?m, while five others sectioned in the horizontal plane had a mean of 38.2 m?m. These neurons produced an average of 6.2 primary dendrites (range: 4–10), many of which projected rostrally or caudally up to 1.0 mm from the cell bodies. These dendrites were oriented along the longitudinal axis; they ran parallel and ventral to the tractus solitarius. In general, all dendrites possessed numerous spines and appendages. Many axons could be traced for considerable distances within the medulla (in one instance, up to 8 mm). These axons were last discerned in the contralateral ventral medulla rostral to the level of their cell bodies. The axons of three neurons bifurcated in the ipsilateral medulla; one branch remained ipsilateral and projected caudally, while the other crossed the midline. A small number of counterstained cells of size similar to or larger than the HRP-stained neurons formed a column that constituted the vl-NTS. Based upon our observations of stained and counterstained cells, we conclude that the inspiratory neurons of the vl-NTS are few in number and represent a morphologically homogeneous population. The primary orientation of the dendritic arbors of vl-NTS inspiratory neurons appears to optimize the surface area available to receive synaptic contacts from sensory afferents emerging from the tractus solitarius.     

Respiration related neurons in the region of the nucleus tractus solitarius of the rabbit

The relationship between external calcium and frequency-facilitated arginine vasopressin (AVP) secretion from the murine neurointermediate lobe was examined in vitro. We evaluated the calcium-dependency of frequency-dependent release in this system, and found that log AVP secretion versus log external calcium plots gave slopes of 0.71, 0.92 and 1.2 for 5, 10 and 20 Hz stimulation, respectively. These slopes are considerably lower than the slopes of 3-4 Hz found at conventional synaptic junctions.     

Synaptic contacts of substance P-immunoreactive axon terminals in the nucleus tractus solitarius onto neurons projecting to the caudal ventrolateral medulla oblongata in the rat

The possibility that substance P (SP)-immunoreactive axon terminals in the nucleus tractus solitarius (NTS) make synaptic contacts onto NTS neurons projecting to the catecholaminergic cell region in the caudal ventrolateral medulla oblongata (CVLM) was examined in the rat using a retrograde tract-tracing method combined with immunohistochemistry. After injection of a retrograde tracer, wheat germ agglutinin-conjugated horseradish peroxidase-colloidal gold complex (WGA-HRP-gold), into the CVLM region where tyrosine hydroxylase-immunoreactive neurons were situated, many retrogradely labeled neurons were detected in the dorsal parts of the NTS, especially at levels between 1.0 mm caudal and 0.5 mm rostral to the obex. Immunoelectron microscopy revealed synaptic contacts between SP-immunoreactive axon terminals and WGA-HRP-gold-labeled neurons in the NTS. These findings indicated that SP regulates NTS neurons which project to the catecholaminergic cell region of the CVLM. ©1997 Elsevier Science B.V. All rights reserved.     

Gustatory functions of the nucleus tractus solitarius in the rabbit

Multiunit analysis revealed a rostral region of NTS containing cells responsive to taste stimulation of rostral tongue. Using representative stimuli for the 4 basic types of taste, maximal incidence and magnitude of response was found to NaCl, followed by HCl, sucrose and QHC1. Further analysis of temporal patterns of response to the tastants revealed differences among stimuli in latency and time course. A principal components analysis indicated that time course, apart from magnitude of response, could contribute to neural differentiation of tastants. Information was also derived on neural intensity functions for these basic types of tastants. Additional observations with sodium saccharin revealed minimal neuronal responsivity despite reported evidence of behavioral preference for this tastant by rabbits.     

Synaptology of the direct projections from the nucleus of the solitary tract to pharyngeal motoneurons in the nucleus ambiguus of the rat

During the pharyngeal phase of the swallowing reflex, the nucleus of the solitary tract (NTS) receives peripheral inputs from the pharynx by means of the glossopharyngeal ganglion and is the location of premotor neurons for the pharyngeal (PH) motoneurons. The semicompact formation of the nucleus ambiguus (AmS) is composed of small and medium-sized neurons that do not project to the pharynx, and large PH motoneurons. We investigated whether the neurons in the NTS projected directly to the PH motoneurons or to the other kinds of neurons in the AmS by using the electron microscope. When wheat germ agglutinin-conjugated horseradish peroxidase (WGA-HRP) was injected into the NTS after cholera toxin subunit B-conjugated HRP (CT-HRP) injections into the pharyngeal muscles of male Sprague-Dawley rats, many nerve terminals anterogradely labeled with WGA-HRP were found to contact PH motoneurons retrogradely labeled with CT-HRP. Most of the labeled axodendritic terminals (63%) contained pleomorphic vesicles with symmetric synaptic contacts (Gray's type II), and the remaining ones contained round vesicles with asymmetric synaptic contacts (Gray's type I). About 14% of the axosomatic terminals on PH motoneuron in a sectional plane were anterogradely labeled, and about 70% of the labeled axosomatic terminals were Gray's type II. Observations of serial ultrathin sections revealed that both the small and the medium-sized neurons received only a few labeled axosomatic terminals that were exclusively Gray's type I. These results indicate that the NTS neurons may send mainly inhibitory as well as a few excitatory inputs directly to the PH motoneurons in the AmS. J. Comp. Neurol. 393:391–401, 1998. © 1998 Wiley-Liss, Inc.     

Cholinergic, nitric oxidergic innervation in cerebral arteries of the cat

Using immunoperoxidase labeling (IPL) and immunofluorescence labeling (IFL) methods, and each followed by NADPH diaphorase (NADPHd) histochemical staining in the same specimen, colocalization of choline acetyltransferase (ChAT) and NADPHd, indicative of nitric oxide synthase (NOS), in cerebral pial arteries and the sphenopalatine ganglia (SPG) of the cat was examined. In addition, retrograde axonal tracing using true blue was performed to determine if cerebral perivascular nerves containing ChAT and NADPHd originate in the SPG. Consistent results were obtained from IPL and IFL methods, indicating that the middle cerebral artery (MCA) and the circle of Willis received dense ChAT-immunoreactive (I) and NADPHd bundles and fine fibers. Almost all ChAT-I fibers and NADPHd fibers were found to be coincident in the arteries examined. A few fine fibers exhibited only NADPHd staining. In the SPG, approximately half of the ganglionic cells were both ChAT-I and NADPHd positive, while the remaining cells were positively only for NADPHd staining. One week after application of true blue on the middle cerebral arteries (MCA), the fluorescent true blue was found in the ganglionic cells of the SPG. Some of the true blue-positive cells contained both ChAT-immunoreactivity and NADPHd staining. These results provide morphological evidence indicating that all ChAT-I fibers in the MCA and the circle of Willis contain NOS, and that these fibers originate in the SPG, although not all NOS-I ganglionic cells in the SPG send fibers to pial vessels. These results also support the hypothesis that acetylcholine (ACh) and nitric oxide (NO) are synthesized and co-released in the same neurons in cerebral perivascular nerves. Based on the reported findings that NO mediates a major component of neurogenic vasodilation, and that ACh acts as a modulator, the present results demonstrate the presence of a cholinergic, nitric oxidergic innervation in cerebral arteries of the cat.     

The ventrolateral medulla as a source of synaptic drive to rhythmically firing neurons in the cardiovascular nucleus tractus solitarius of the rat

We sought to determine whether the caudal ventrolateral medulla (cVLM), at the level of area postrema, influences the rhythmically beating neurons found within the dorsomedial NTS in rat brainstem slices. Intra- or extracellular recordings of neurons firing rhythmically at around 5 Hz were characterized as either auto-active (i.e. pacemaker; AA) or synaptically driven (SD) by pharmacological interventions. The nature of inputs evoked from the ipsilateral cVLM were orthodromic and the majority were excitatory (latency 3-20 ms). Further, this excitatory influence was found to be tonically active in 25/47 cells studied since inactivating the ipsilateral cVLM by localized cooling reduced the firing rate by 0.5-3.0 Hz (23% on average). Neuronal characterization showed that the most consistent and pronounced effect occurred on SD rather than AA cells. Control experiments that cooled other areas of the slice closer to the recording site proved ineffective. Additional studies showed that most rhythmically firing cells in the NTS received an excitatory synaptic input from the solitary tract (ts; latency 3-30 ms). This input was reduced or blocked by inactivating the cVLM in neurons in which the ts latency of activation was greater than 8 ms in half of the neurons tested. Subsequent pharmacological tests revealed that these neurons were predominantly SD. Identified AA neurons received an input from the ts at a shorter latency, typically less than 8 ms, and this was unperturbed by cooling the cVLM in all cases. Further, there was no obvious difference in the baseline discharge rates between cells in the hemi-slice and those recorded in an intact slice. In a hemi-coronal slice cooling the cVLM also produced a 20% decrease in firing rate in identified SD neurons but no consistent change in AA cells. We conclude that (1) the ipsilateral cVLM contributes principally tonic excitatory drive to rhythmically active neurons in the dorsomedial NTS in vitro and this preferentially effects SD neurons; (2) other excitatory drives other than those from the ipsilateral cVLM impinge upon SD cells, the origin of which are relatively local and likely to be in the NTS; (3) in the slice the projection from the cVLM to the NTS appears to be present but the reciprocal connection is absent.     

Morphology of physiologically identified slowly adapting lung stretch receptor afferents stained with intra-axonal horseradish peroxidase in the nucleus of the tractus solitarius of the cat. II. An ultrastructural analysis

The nucleus of the tractus solitarius is a site for termination of primary afferents originating from a variety of visceral receptors. The localization of bouton terminals of slowly adapting lung stretch (SAR) afferent fibers originating from the tracheobronchial tree have been described in the companion paper (Kalia and Richter, '85). The most conspicuous finding regarding the location of SAR terminals is that they are concentrated within specific subnuclear groups of the nucleus of the tractus solitarius (nTS) and are distributed widely in the rostrocaudal plane of the medulla oblongata. These light microscopic features have provided us with valuable information with regard to the organization of visceral afferents in the central nervous system. The synaptic profiles formed by the 476 bouton terminals of these HRP-labeled afferents have been described in this paper in serial thin sections. All of the bouton terminals examined under the electron microscope were found to contain round synaptic vesicles. Synaptic boutons (1.0-3.0 microns in diameter) were usually of the en passant variety and made contact with different structures depending upon the subnucleus which was examined. In the ventral (v) and the ventrolateral (vl) subnuclei of the nTS, asymmetrical (type I) synaptic contacts containing round, clear synaptic vesicles of 35-50 microns in diameter were found and these contacts were made with (1) the soma of cell bodies located in that subnucleus; (2) spiny dendrites in that nucleus; (3) vesicle-containing axon terminals that were presynaptic to the HRP-labeled bouton terminal; and (4) vesicle-containing dendrites in which the HRP profile was presynaptically located. The terminal axon remained myelinated till the last 1 micron before the bouton terminal was formed. There was no distinct, unmyelinated portion of the terminal axon. The synaptic bouton received axon-axonal synapses from unlabeled bouton terminals containing round, clear vesicles. This is the first report of the localization of these afferent fibers as well as of the regional variations in the ultrastructure of boutons of physiologically identified terminals. It appears likely that the lung stretch afferent fibers, by having axon-axonal as well as axon-somatic contact in the ventral, ventrolateral, and intermediate subnuclei of the nTS, can interact in a variety of different ways in this region. The significance of these features in relation to the precise influence of respiratory afferents on central respiratory mechanisms needs to be evaluated further.     

Synaptic responses of neurons of the nucleus tractus solitarius in vitro

Postsynaptic responses of neurons in the nucleus tractus solitarius (NTS) have been studied in an in vitro slice preparation using extra- and intracellular recording. Single or paired pulse stimulations were delivered to afferent fibers within the tractus solitarius (TS) to activate orthodromic responses in these neurons. Most NTS neurons displayed an initial synaptic excitation followed by inhibition of spontaneous or evoked firing lasting up to 150-200 ms after stimulation. Excitatory postsynaptic potentials (EPSPs), recorded intracellularly, were increased in amplitude by membrane hyperpolarization. Large afterhyperpolarizations followed action potentials triggered by the EPSPs or evoked by intracellular current injections. Intracellular evidence for synaptic inhibition within the NTS included: (1) the presence, after Cl-injection, of flurries of spontaneous PSPs likely to be inverted inhibitory postsynaptic potentials; (2) reduction of the size of a test EPSP by a previous subthreshold TS conditioning volley; and (3) hyperpolarizing PSPs recorded in some neurons. Other NTS neurons exhibited prolonged excitatory responses to TS stimulation and could be local inhibitory interneurons. These results may help specify synaptic mechanisms in the NTS that could play an integrative role in the relay of visceral sensory inputs to higher order effectors.     

Multimodal responses of taste neurons in the frog nucleus tractus solitarius

The responses of 216 neurons in the nucleus tractus solitarius (NTS) of the American bullfrog were recorded following taste, temperature, and tactile stimulation. Cells were classified on the basis of their responses to 5 taste stimuli: 0.5 M NaCl, 0.0005 M quinine-HCl (QHCl), 0.01 M acetic acid, 0.5 M sucrose, and deionized water (water). Neurons showing excitatory responses to 1, 2, 3, or 4 of the 5 kinds of taste stimuli were named Type I, II, III, or IV, respectively. Cells whose spontaneous rate was inhibited by taste and/or tactile stimulation of the tongue were termed Type V. Type VI neurons were excited by tactile stimulation alone. Of the 216 cells, 115 were excited or inhibited by taste stimuli (Types I-V), with 35 being Type I, 34 Type II, 40 Type III, 2 Type IV and 4 Type V. The remaining 101 cells were responsive only to tactile stimulation (Type VI). Of those 111 cells excited by taste stimulation (Types I-IV), 106 (95%) responded to NaCl, 66 (59%) to acetic acid, 44 (40%) to QHCl, 10 (9%) to water, and 9 (8%) to warming. No cells responded to sucrose. Of the 111 cells of Types I-IV, 76 (68%) were also sensitive to mechanical stimulation of the tongue. There was some differential distribution of these neuron types within the NTS, with more narrowly tuned cells (Type I) being located more dorsally in the nucleus than the more broadly tuned (Type III) neurons. Cells responding exclusively to touch (Type VI) were also more dorsally situated than those responding to two or more taste stimuli (Types II and III).     

Antidromic mapping of descending axons of respiratory bulbospinal neurons in the nucleus tractus solitarius of the rabbit

Antidromic mapping of the descending axons of the respiratory bulbospinal neurons in the region of the nucleus tractus solitarius (NTS) was performed on rabbits anesthetized with urethane. Among 177 units tested, 29 out of 87 inspiratory (I), 27 out of 84 expiratory (E) and 2 out of 6 phase-spanning units were identified as bulbospinal. A prominent of the bulbospinal pathway from the NTS in the rabbit is the abundance of ipsilateral descending axons. The axons rising from one side are situated in the ventrolateral and ventral funiculi of both sides. The axonal conduction velocities are about 25–35 m/s. Both I and E bulbospinal neurons can be divided into R_α and R_β types according to ‘no I inflation’ and ‘maintained E inflation’ tests.     

Inhibitory effect of cholecystokinin octapeptide on neurons in the nucleus tractus solitarius

We investigated the effect of the cholecystokinin octapeptide (CCK8) applied locally to neurons of the nucleus tractus solitarius (NTS). Results demonstrate an inhibitory effect of CCK8 on spike discharges including those related to respiration. It is suggested that CCK8 acts at this level through specific receptor mechanisms since CCK8-induced inhibitions were not reproduced by application of related peptides and were resistant to antagonists of different inhibitory transmitters.