Influence of rubrospinal tract and the adjacent mesencephalic reticular formation on the activity of medullary respiratory neurons and the phrenic nerve discharge in the rabbit

Suprapontine brain sites acting on the central respiratory system have been demonstrated to give rise to inspiratory as well as expiratory facilitatory effects. In the present study the inspiratory inhibitory effect which has been reported in the cat to be elicited consistently by electrical stimulation of the rubrospinal tract and the adjacent mesencephalic reticular formation was examined in the urethane-anaesthetized rabbit. Stimulation of these sites with single electrical shocks of moderate intensity induced a short latency (onset after 3.0 ms) transient (duration: 29 ms) inhibition of the phrenic nerve activity (PHR). Short volleys of stimuli applied in mid- to late-inspiration led to a premature off-switch of inspiration. The extracellularly recorded discharge activity of the different types of medullary respiration-related units (RRU) reflected these alterations, accordingly. Axonal connections of RRU with mesencephalic structures were evaluated. Examination of orthodromic responses of medullary RRU to stimulation of this pathway revealed that most bulbospinal inspiratory neurons (10 out of 13) were paucisynaptically inhibited after short latency (at least 1.2 ms). The conduction time from bulbospinal inspiratory neurons to the recording site of PHR was 1.6 ms. Thus, a disynaptic pathway — including bulbospinal inspiratory neurons — is suggested inducing inspiratory inhibition 3.0 ms after single shock midbrain stimulation. This inhibition results in disfacilitation of phrenic motoneurons. The fact that extensive electrolytic lesions of the pneumotaxic center in rostral pons did not abolish the observed inspiratory inhibitions excludes these structures from being involved. A direct pathway from the red nucleus and the adjacent reticular formation to phrenic nuclei of the spinal cord, however, can not be excluded from being involved in the demonstrated inspiratory inhibition. The described effects may play a role in behavioral or voluntary control of respiration.     

Respiratory-modulated neuronal activities of the rostral medulla which may generate gasping

Different neurophysiological mechanisms have been proposed to generate eupnea and gasping. Gasping is generated by neuronal mechanisms intrinsic to the medulla whereas a ponto-medullary neuronal circuit has been hypothesized to generate eupnea. Hence, neurons in the rostral medullary region which are critical for the neurogenesis of gasping are hypothesized to discharge differently in eupnea and gasping. In a perfused in situ preparation of the juvenile rat, these rostral medullary neuronal activities had inspiratory, expiratory and phase-spanning patterns in eupnea. In gasping, most expiratory and phase-spanning activities ceased, whereas many inspiratory neuronal activities changed to a decrementing pattern as that of the phrenic nerve. A limited proportion of neuronal activities acquired a 'pre-inspiratory' discharge in gasping. These neuronal activities, which were inspiratory or phase-spanning in eupnea, commenced discharge in neural expiration. This discharge peaked at the onset of the gasp and then decremented during neural inspiration. We hypothesize that these 'pre-inspiratory' neuronal activities generate the gasp by intrinsic pacemaker mechanisms.     

Localization and properties of respiratory neurons in the rostral pons of the newborn rat

The distribution and discharge pattern of respiratory neurons in the ‘pneumotaxic center’ of the rostral pons in the rat has remained unknown. We performed optical recordings and whole-cell patch clamp recordings to clarify respiratory neuron activity in the rostral pons of a brainstem-spinal cord preparation from a newborn rat. Inspiratory nerve activity was recorded in the 4th cervical nerve and used as a trigger signal for optical recordings. Respiratory neuron activity was detected in the limited region of the rostral-lateral pons. The main active region was presumed to be primarily the Kölliker-Fuse nucleus. The location of respiratory neurons was further confirmed by Lucifer Yellow staining after conducting whole-cell recordings. From a membrane potential analysis of the respiratory neurons in the rostral pons, the respiratory neurons were divided into four types: inspiratory neuron (71.9%), pre-inspiratory neuron (5.3%), post-inspiratory neuron (19.3%), and expiratory neuron (3.5%). A noticeable difference between pontine and medullary respiratory neurons was that post-inspiratory neurons were more frequently encountered in the pons. Application of a μ-opioid agonist, [d-Ala2, N-Me-Phe4, Gly5-ol]-enkephalin, transformed the burst pattern of post-inspiratory neurons into that of pre-inspiratory neurons. The electrical stimulation of the sensory root of the trigeminal nerve induced three types of responses in 85% of pontine respiratory neurons: inhibitory postsynaptic potentials (42.7%), excitatory postsynaptic potentials (37.7%) and no response (15.1%). Our findings provide the first evidence in the rat for the presence of respiratory neurons in the rostral pons, with localization in the lateral region approximately overlapping with the Kölliker-Fuse nucleus.     

Excitatory and inhibitory synaptic inputs shape the discharge pattern of pump neurons of the nucleus tractus solitarii in the rat

The second-order relay neurons of the slowly-adapting pulmonary stretch receptors (SARs) are called pump neurons (P cells) and are located in the nucleus tractus solitarii (NTS). We have shown recently that P cells do not act merely as simple relay neurons of SAR afferents but also receive rhythmic inputs from the central respiratory system. This study aimed to analyze two aspects of the respiratory inputs to P cells: (1) suppression of P cell firing at early inspiration (eI suppression) and (2) facilitation of P cell firing at around the period from late inspiration to early expiration (IE facilitation). This study employed extracellular recordings combined with iontophoretic applications of neuroactive drugs to single P cells, in Nembutal-anesthetized, paralyzed, and artificially ventilated rats. The results showed that several excitatory and inhibitory neurotransmitters were involved in these synaptic events. First, the glycine antagonist strychnine and the GABA_A antagonist bicuculline were applied to identify the neurotransmitters acting in eI suppression. Strychnine greatly diminished eI suppression, but bicuculline had little effect. This suggested that eI suppression was elicited by inspiratory neurons that were glycinergic and had a decrementing firing pattern. Second, on the other hand bicuculline markedly enhanced IE facilitation as well as the baseline frequency of P cell firing. The enhancement of IE facilitation was distinctive even when the effects of increased baseline firing on this enhancement were taken into account. Third, IE facilitation was diminished by applications of the NMDA glutamate receptor antagonists D-2-amino-5-phosphonovaleric acid (APV) and dizocilpine (MK-801). These results suggested that glutamatergic synapses on P cells from some unidentified respiratory neurons form excitatory inputs for IE facilitation and GABA_A receptor-mediated processes control the strength of IE facilitation, possibly at the presynaptic level. Finally, iontophoretic application of the non-NMDA glutamate receptor antagonist, 6-cyano-7-nitroquinoxaline-2, 3-dione disodium (CNQX), almost completely abolished P cell firing in response to both lung inflation and electrical stimulation of the vagus nerve. This confirmed the previous report that glutamate is the primary neurotransmitter at the synapses between SAR afferents and P cells. We concluded that complicated synaptic inputs involving glycinergic and GABAergic inhibitions, and non-NMDA and NMDA glutamate receptor-mediated excitations form the basic pattern of P cell firing. Received: 31 March 1999 / Accepted: 8 June 1999     

Descending axonal projections from the medial parabrachial and K?lliker-Fuse nuclear complex to the nucleus raphe magnus in cats.

In 9 Nembutal-anesthetized and vagotomized cats, a total of 42 units, including 2 respiratory units, recorded from the medial parabrachial (NPBM) and K?lliker-Fuse (KF) nuclear complex were found to be antidromically activated by electrical stimulation of the nucleus raphe magnus (NRM). The latencies ranged from 0.4 to 2.5 ms (mean 1.1 ms). In 5 cats, following injection of WGA-HRP (wheat germ agglutinin-conjugated horseradish peroxidase) into the NRM, a number of retrogradely labelled neurons were observed in the rostral pons, mainly in the NPBM, KF and nearby pontine area. These results demonstrate that mainly non-respiratory neurons in the rostral pons, especially in the NPBM and KF nucleus, send monosynaptic axonal projections to the NRM.     

Inhibition of medullary reticulospinal neurons by excitation of the dorsolateral parts of the pons which block movement and muscle tone in rats

Analysis of the response of 128 reticulospinal neurons in the magnocellular and ventral reticular nuclei showed that 36.7% of these cells responded with short-latency (2–4 msec) action potentials and increased their tonic activity in response to electrical stimulation of the central parts of the hypothalamus, which evoked increases in hindlimb muscle tone in rats. These cells completely stopped producing action potentials during electrical stimulation and during chemical stimulation of the dorsolateral parts of the pons, which inhibited movement and muscle tone. A total of 23.4% of the cells produced only short-latency (1–4 msec) action potentials in response to stimulation of the inhibitory parts of the pons. A total of 3.9% of reticulospinal neurons increased their activity during stimulation of the hypothalamic zones and pontine areas of the brain. No responses were obtained from 35.9% of neurons. It is suggested that excitation of pontine structures inhibiting movement and muscle tone may prevent conduction in descending activatory systems from the rostral parts of the brain (which increase muscle tone) to the reticulospinal neurons of the medulla oblongata. Translated from Rossiiskii Fiziologicheskii Zhurnal imeni I. M. Sechenova, Vol. 85, No. 3, pp. 353–359, March, 1999.     

Olfactory and visceral projections to the nucleus of the solitary tract

Electrophysiological studies were performed to determine if neurons of the nucleus of the solitary tract (NTS) which receive inputs from the stomach via vagal afferents also respond to olfactory bulb (OB) stimulation. The frequency of neuronal activity of the rostral ventral portion of the NTS was increased by gastric distension (GD). The evoked potentials in the same site due to vagal stimulation displayed short latencies; whereas, the evoked potentials in the dorsomedial part of the NTS due to vagal stimulation had considerably longer latencies. Gastric distension decreased neuronal activity in the dorsomedial NTS. Evoked potentials and increases in neuronal activity were also observed in the dorsomedial NTS due to electrical stimulation. In the dorsomedial NTS, OB stimulation enhanced the decrease in neuronal activity due to GD. Olfactory and visceral functions apparently interact in the NTS in modulating taste mechanisms involved in food selection and ingestion.     

Convergence of cortico- and cuneopontine projections onto components of the pontocerebellar system in the rat: an anatomical and electrophysiological study

Summary Previous studies in the rat have demonstrated that corresponding peripheral tactile and somatosensory cortical inputs converge within the granule cell layer of various cerebellar lobules and further that descending corticopontine projections from the forelimb sensory cortex (FLSCx) partially overlap with the projection zones of ascending basilar pontine afferents from nucleus cuneatus (NC). The present study employed anatomical and electrophysiological procedures to determine whether cortical and dorsal column nuclear afferent projections converge on pontine neurons that, in turn, provide mossy fiber input to the granule cell layer of the paramedian lobule (PML), i. e., that portion of the rodent cerebellum shown to receive forelimb peripheral inputs. The combination of the orthograde and retrograde axonal transport of horseradish peroxidase (HRP) conjugated to wheat germ agglutinin (WGA) was used light microscopically to demonstrate that orthogradely labeled projections from injections of the FLSCx and NC converged with ponto-paramedian projection neurons that were retrogradely labeled from injections of the PML. These studies were also repeated in conjunction with ablations of either the FLSCx or NC which resulted in the ultrastructural identification of degenerating, as well as WGA-HRP labeled axonal boutons of these pontine afferent projections thus confirming that such projections actually formed synaptic contacts with the retrogradely labeled pontoparamedian projection neurons. Single unit recording analyses of neurons in the ventromedial region of the basilar pons following combined electrical stimulation of various regions of the sensorimotor cortex and the contralateral body surface indicated that approximately 40% of all cells recorded responded to electrical stimulation of corresponding regions of the cortex and periphery, particularly the FLSCx and the forepaw. Natural cutaneous stimuli applied to the forepaw that also elicited responses in these same groups of basilar pontine neurons and were associated with relatively small receptive fields. Taken together, these observations indicate that the previously observed convergence of peripheral and somatosensory cortical inputs within the granule cell layer of the cerebellar cortex may be at least partially organized at the level of the basilar pons.     

Electrical stimulation of arterial and central chemosensory afferents at different times in the respiratory cycle of the cat: II. Responses of respiratory muscles and their motor nerves

The response patterns of the electrical activity of the respiratory motor nerves and muscles to brief electrical stimulation of the arterial and the intracranial chemosensory afferents were studied in anesthetized cats. Stimulation during inspiration increased the activity of phrenic nerve and the inspiratory muscles (intercostal, diaphragm) with a latency of 15–25 ms, whereas expiratory muscle activity in the following expiration remained almost unaltered. Stimulation during expiration increased the activity of expiratory nerves and muscles (intercostal, abdominal) after a delay of 80–120 ms. The later the stimulation occurred in the insor expiratory period the larger the increase in amplitude and in steepness of rise of the respective integrated activity in respiratory nerves and muscles. Stimulation in early inspiration shortened the discharge period of inspiratory muscles, whereas excitation in early expiration caused an earlier onset and prolonged the activity in the expiratory muscles. Stimulation in the late phase of ins- or expiration prolonged the discharge of the respective nerves and muscles. Both the arterial (carotid sinus nerve, CSN, and aortic nerve, AN) and intracranial chemosensory (VM) afferents stimuli were able to affect both the inspiratory and the expiratory mechanisms. The restriction of the effects to the phase of the stimulus suggests a mechanism by which these afferents, when activated during inspiration, effectively project only to inspiratory neurones, and vice versa for expiration.Supported by the Deutsche Forschungsgemeinschaft, SFB 114 Bionach     

Pontine respiratory group neuron discharge is altered during fictive cough in the decerebrate cat

A network of neurons in the rostral dorsal lateral pons and pons/mescencephalic junction constitute the pontine respiratory group (PRG) and is essential for reflex cough. As a next step in understanding the role of the PRG in the expression of the cough reflex, we examined neuron firing rates during fictive cough in cats. Decerebrated, thoracotomized, paralyzed, cycle-triggered ventilated adult cats were used. Extracellular activity of many single neurons and phrenic and lumbar neurograms were monitored during fictive cough produced by mechanical stimulation of the intrathoracic trachea. Neurons were tested during control periods for respiratory modulation of firing rate by cycle-triggered histograms and statistical tests. Most respiratory modulated cells were continuously active with various superimposed respiratory patterns; major categories included inspiratory decrementing (I-Dec), expiratory decrementing (E-Dec) and expiratory augmenting (E-Aug). There were alterations in the discharge patterns of respiratory, as well as, non-respiratory modulated neurons during cough. The results suggest an involvement of the PRG in the configuration of the cough motor pattern.     

The pontocerebellar projection to the uvula in the cat

Summary The occurrence of retrogradely labeled cells in the pontine nuclei was mapped following injections of 0.3–0.4 l of a horseradish peroxidase suspension (50% weight/volume) into the uvula (lob. IX of Larsell) in the cat. The uvula was found to receive afferents from three pontine cell collections. One of these is situated in the paramedian pontine nucleus close to the midline. It forms a fairly distinctly outlined longitudinal column of cells and is present at all levels of the pons except most rostrally and caudally. Another group, in the dorsolateral and lateral pontine nuclei, extends as a somewhat shorter cell column in the longitudinal direction. The third region consists of cells within the rostral part of the peduncular nucleus in its dorsomedial region. The pontine projection to the uvula is bilateral, with some preponderance of crossed connections.The projection to the uvula is organized according to the pattern determined previously for pontine projections to other parts of the cerebellum. A single lobule or part of it receives afferents from more than one cell group in the pons. The projecting cells are most often arranged in longitudinal columns.Correlations with data on the termination of afferents to the pons permit some conclusions regarding the sources of information reaching the uvula via the pons. Main sources seem to be the superior and inferior colliculi, the intracerebellar nuclei and the sensorimotor cortices.     

Synaptic interactions between respiratory neurons during inspiratory on-switching evoked by vagal stimulation in decerebrate cats

To elucidate neuronal mechanisms underlying phase-switching from expiration to inspiration, or inspiratory on-switching (IonS), postsynaptic potentials (PSPs) of bulbar respiratory neurons together with phrenic nerve discharges were recorded during IonS evoked by vagal stimulation in decerebrate and vagotomized cats. A single shock stimulation of the vagus nerve applied at late-expiration developed an inspiratory discharge in the phrenic neurogram after a latency of 79+/-11 ms (n = 11). Preceding this evoked inspiratory discharge, a triphasic response was induced, consisting of an early silence (phase 1 silence), a transient burst discharge (phase 2 discharge) and a late pause (phase 3 pause). During phase 1 silence, IPSPs occurred in augmenting inspiratory (aug-I) and expiratory (E2) neurons, and EPSPs in postinspiratory (PI) neurons. During phase 2 discharge, EPSPs arose in aug-I neurons and IPSPs in PI and E2 neurons. These initial biphasic PSPs were comparable with those during inspiratory off-switching evoked by the same stimulation applied at late-inspiration. In both on- and off-switching, phase-transition in respiratory neuronal activities started to arise concomitantly with the phrenic phase 3 pause. These results suggest that vagal inputs initially produce a non-specific, biphasic response in bulbar respiratory neurons, which consecutively activates a more specific process connected to IonS.     

Stimulation of the pedunculopontine tegmental nucleus in the rat produces burst firing in A9 dopaminergic neurons.

Stimulation of the medial prefrontal cortex in the rat produces events in midbrain dopaminergic neurons which resemble natural bursts, and which are closely time-locked to the stimulation, albeit with a very long latency. As a consequence, we have previously argued that such bursts are polysynaptically generated via more proximal excitatory amino acidergic afferents, arising, for example, from the pedunculopontine tegmental nucleus. In the present study, single-pulse electrical stimulation applied to this nucleus (and other sites in the rostral pons) was found to elicit responses in the majority of substantia nigra (A9) dopaminergic neurons. Responses usually consisted of long-latency, long-duration excitations or inhibition-excitations. Thirty-seven percent of responses (currents combined) elicited by stimulation of the pedunculopontine tegmental nucleus contained time-locked bursts, the bursts being embedded in the long-duration excitatory phases of excitation and inhibition-excitation responses. Stimulation sites located within 0.5 mm of the pedunculopontine tegmental nucleus were also effective at eliciting time-locked bursts (although less so than sites located in the nucleus itself), whereas more distal sites were virtually ineffective. For responses containing time-locked bursts, a higher percentage of stimulations produced a burst when the response was elicited from within the pedunculopontine tegmental nucleus than when it was elicited from outside: the bursts themselves having a very long latency (median of 96.2 ms; shorter than that of medial prefrontal cortex-induced bursts). Finally, although there was no difference in the distribution within the substantia nigra pars compacta of cells which exhibited time-locked bursting and those which did not, stimulation-induced bursts were elicited more frequently in dopaminergic neurons which were classified as "bursting" on the basis of their basal activity. The pedunculopontine tegmental nucleus appears to be a critical locus in the rostral pons for the elicitation of time-locked bursts in A9 dopaminergic neurons. Since time-locked bursts were more often elicited from cells which exhibited bursting under basal conditions, this suggests that rostral pontine sites, in particular the pedunculopontine tegmental nucleus, may play a role in the natural burst activity of dopaminergic neurons. Given that bursts in dopaminergic neurons are generated in response to primary and secondary reinforcers, the projection from the pedunculopontine tegmental nucleus could be one means by which motivationally relevant information (arising, for example, from the medial prefrontal cortex) reaches these cells.     

Effects of stimulation of phrenic afferent fibers on medullary respiratory neurons in cat

The effects of electrical stimulation of both cervical branches (C5 and C6) of the right phrenic nerve on medullary respiratory neuron activity were studied in anesthetized, spontaneously breathing cats. In 14 cats, the stimulation of the thin phrenic afferents had no effect on the inspiratory duration and evoked excitatory or inhibitory responses in only 3/86 inspiratory neurons tested. In 3 cats, the stimulation decreased the inspiratory duration and 26/26 inspiratory neurons showed a shortened discharge without modification of their discharge frequency. Although the effects of the stimulation were not analysed by averaging techniques, it is concluded that phrenic afferents do not exert an important control on the medullary respiratory neuron discharge.     

Vagal modulation of responses elicited by stimulation of the aortic depressor nerve in neurons of the rostral ventrolateral medulla oblongata in the rat.

Stimulation of cervical vagal afferents inhibits central sympathetic outflows in part by inhibiting the ongoing activity of putative baroreceptive neurons in the rostral ventrolateral medulla oblongata. The aim of the present study was to examine the electrophysiological characteristics of vagal responses and their interactions with responses elicited by stimulation of the aortic nerve in neurons there. The study focused on the role of the long-lasting, late-onset vagal inhibition, which is likely to play an important role in the tonic inhibitory effects of vagal afferent stimulation. In vivo intracellular recordings were obtained from 33 neurons that received convergent inputs from aortic and vagal afferents. Sixty-four percent of these neurons exhibited a late inhibition following electrical stimulation of myelinated vagal afferents (mean onset latency of 100+/-5 ms). The average duration of late inhibition (294+/-19 ms) exceeded the duration of the cardiac cycle. As a consequence of this, sustained vagal stimulation diminished the effect of rhythmic baroreceptor inputs in neurons that exhibited late vagal inhibition. Simultaneous activation of aortic and vagal afferents significantly increased the magnitude of late inhibition, even in those neurons where stimulation of the aortic nerve alone did not elicit a response (n = 15). This suggested that the convergence between vagal and aortic afferent inputs occurred in inhibitory inteneurons antecedent to the recorded rostral ventrolateral medulla oblongata neurons. Focal stimulation of the caudal part of the nucleus of the solitary tract also elicited a late-onset inhibition in 73% of the neurons that responded to stimulation of the aortic nerve. This inhibition appeared to be similar to the late vagal inhibition, except for its shorter average onset latency (64+/-7 ms). Based on this observation, it is proposed that inhibitory inteneurons that mediate late inhibition to rostral ventrolateral medulla oblongata neurons may lie within the caudal part of the nucleus of the solitary tract. The present study established that activation of myelinated vagal afferents exerts a complex modulation over the ongoing and evoked activity of neurons that respond to stimulation of the aortic nerve. The complex interaction that occurs between aortic and vagal inputs in neurons of the rostral ventrolateral medulla may be implicated in long-term modulation of sympathetic outflows in response to changes in the activation of visceral receptors supplied by vagus afferents. The modulation elicited by late vagal inhibition may help to adjust cardiovascular outflows according to requirements set by the thoraco-abdominal visceral environment.     

Brain stem reticular formation neuronal correlates of stimulus significance and behavior during discriminative avoidance conditioning in rabbits

Multiple-unit activity in the reticular formation of the midbrain and pons was recorded in rabbits during discriminative conditioning of locomotor (wheel-running) avoidance behavior. The conditional stimuli (CS+ and CS-) were pure tones of different auditory frequency, and the unconditional stimulus (US) was a constant-current footshock (1.5-2.5 mA) delivered through the grid floor of the wheel. The pontine, but not the midbrain, sites manifested development during behavioral acquisition, of brief-latency (10-40-ms) discriminative neuronal discharges (i.e., greater discharges to the CS+ than to the CS-). The greatest magnitude of the discriminative discharges in the rostral pontine loci occurred in the first conditioning session. The discriminative response in the caudal loci developed more slowly, and it persisted to the criterial stage of training. Both rostral and caudal pontine loci, during the interval from CS onset to US onset, manifested a progressive build-up of neuronal firing in anticipation of the behavioral response. The occurrence in the rostral and caudal pontine areas, respectively, of early- and late-developing discriminative discharges is analogous to effects observed in past studies in the limbic mesocortical and thalamic systems. These findings provide a basis for establishing the possible functional relatedness or independence of these analogous effects.     

Glutamate immunoreactivity in the rat basilar pons: light and electron microscopy reveals labeled boutons and cells of origin of afferent projections.

Immunohistochemical methods that employed a polyclonal antiserum directed against a glutamate-hemocyanin conjugate were utilized to examine the rat basilar pontine nuclei at both light and electron microscopic levels in order to identify putative glutamatergic neural elements. A large number of cells ranging in size from 11 to 32 microns in diameter and present in all subdivisions and at all rostrocaudal levels of the basilar pons exhibited intense glutamate immunoreactivity. Immunoreactive punctate structures, confirmed by electron microscopy to be axon terminals, were homogeneously distributed throughout the pontine neuropil, although a somewhat greater accumulation was apparent medially at mid-levels of the basilar pons and laterally at more caudal levels. Immunolabeled axons were also present throughout the pontine nuclei. In order to demonstrate possible extrinsic sources of glutamate-immunoreactive axon terminals within the pontine gray, injections of wheat germ agglutinin-horseradish peroxidase were made directly into the basilar pons. Tissue was then evaluated for the presence of retrogradely transported wheat germ agglutinin-horseradish peroxidase and the same tissue sections processed for glutamate immunocytochemistry. Following this combined protocol, neuronal somata exhibiting both wheat germ agglutinin-horseradish peroxidase and glutamate immunoperoxidase reaction products were observed within layer Vb of the cerebral cortex, zona incerta, the dentate nucleus of the cerebellum, nucleus paragigantocellularis of the medullary reticular formation, and the dorsal column nuclei. Such double-labeled cells were considered to represent glutamatergic neurons that provide axonal projections to the basilar pons. Ultrastructural studies of the pontine nuclei confirmed the presence of glutamate immunogold labeling in dendrites, neuronal somata, axons, and axon terminals. Immunoreactive boutons contained round vesicles and primarily formed asymmetric synapses at various postsynaptic loci which included glutamate-immunolabeled dendritic profiles and somata. These results suggest that glutamatergic basilar pontine neurons form one segment of a multisynaptic pathway involving glutamatergic afferents to the basilar pons, glutamatergic pontocerebellar projection neurons, and the glutamatergic granule cells of the cerebellar cortex.     

Evidence for a respiration-modulated cholinergic action on the activity of medullary respiration-related neurons in the rabbit

Effects of the iontophoretically administered cholinergic agonists acetylcholine, bethanechol and DMPP on the activity of medullary respiration-related neurons were examined in urethane-anaesthetized rabbits. Inhibitory effects prevailed over excitatory effects. Analysis of cholinergic effects by cycle-triggered averaging revealed three major types of neuronal responses: (i) constant alterations of spike-density throughout the whole period of activity ("constant effects"), (ii) effects increasing during the progression of the burst of discharge or effects restricted to a particular fraction of the burst ("phasic effects") and (iii) effects which were characterized by an excitation during one respiratory phase and an inhibition during the other phase ("bi-phasic effects"). The latter type of effects was observed in phase-spanning respiration-related neurons. Phasic effects were mainly observed in inspiration-related neurons which were predominantly inhibited by stimulation of muscarinic receptors. Inspiratory R beta-neurons in no case were phasically affected by cholinergic agents. The mean muscarinic inhibition of inspiration-related neurons increased with the progression of inspiration. The mean nicotinic inhibition of expiration-related neurons decreased with the progression of expiration. Results suggest that the efficacy of (i) a central inspiration terminating mechanism and (ii) the onset of discharge of expiratory neurons is modulated by acetylcholine.     

The pontocerebellar projection in the rhesus monkey: an experimental study with retrograde axonal transport of horseradish peroxidase.

The pontocerebellar projection has been studied in the Rhesus monkey by use of the retrograde axonal transport of horseradish peroxidase. The parts of the cerebellum investigated receive afferents from pontine cell groups arranged as rostro-caudally oriented lamella or slab-like regions. As a rule labelled cells are found at all rostro-caudal levels in discrete groups, but their number at various levels, and particularly their distribution in the transverse plane, differ according to which part of the cerebellum has been injected. Although the same cell group may apparently be labelled after injections of different parts of the cerebellum, each cerebellar region has its own characteristic territory in the pontine nuclei. The anterior lobe receives fibres from lamella-like regions mainly in the caudal half of the pons, with cells projecting to the vermis and intermediate parts of the anterior lobe somewhat differently situated. Crus I is connected mainly with a region rostromedially in the pons, while crus II receives fibres from all levels of the pons, the cells being located medially and ventrally at caudal levels and more laterally at rostral levels. The paramedian lobule is supplied from pontine cells restricted to the rostral two-thirds of the pons and located more laterally than those projecting to crus II. Lobules VII and VIIIA (the main part of the vermal visual area) receive fibres mainly from two long column-like regions located dorsomedially and dorsolaterally, while lobulus VIIIB is supplied mainly from other areas in the pons.Counting of labelled cells indicates that about 30% of the fibres to the vermis, and about 10% to the hemispheres, are uncrossed. The density of the projection to the cerebellar hemispheres is found to be about three times as high as to the vermis. In two cases injections of horseradish peroxidase were placed in the cerebral cortex as well as in the cerebellum, enabling a direct comparison of sites of termination of corticopontine fibres (orthograde transport of horseradish peroxidase) with location of pontocerebellar neurons.It is suggested that the cortico-ponto-cerebellar pathway is organized so as to bring about convergence in the cerebellar cortex from several parts of the cerebral cortex, and that this takes place in a highly organized manner so that each small part of the cerebellar cortex has its own characteristic set of inputs.     

Peripheral chemoreceptor inputs to medullary inspiratory and postinspiratory neurons of cats

The effect of peripheral chemoreceptor activation on inspiratory and postinspiratory medullary neurons was investigated using intracellular recording techniques. Peripheral chemoreceptors were activated by injecting CO₂ saturated 1 N bicarbonate solution into the lingual artery or by electrically stimulating the carotid sinus nerve. Injections of 20–300 l bicarbonate solution evoked changes in respiratory frequency and in peak phrenic nerve discharge. The membrane potential of inspiratory alpha neurons, whether bulbospinal or not and independent of their anatomic location, was decreased during inspiration. A sequence of compound excitatory and inhibitory effects were observed when the stimulus was given during the postinspiratory and expiratory phases of the respiratory cycle. Inspiratory beta- and late-inspiratory neurons, however, were inhibited by peripheral chemoreceptor activation. Postinspiratory neurons were strongly activated during postinspiration. Neither class of respiratory neurons were shown to receive direct synaptic inputs from the peripheral chemoreceptors as tested by electrical stimulation of the carotid sinus nerve and signal averaging of the respiratory neuron membrane potential. The experiments revealed differential influences of afferent chemoreceptor activity on various components of the respiratory network. We conclude that chemoreceptor afferents activate non-respiratory modulated medullary neurons which, in turn, activate or inhibit various neurons of the medullary respiratory control network. The responses of each type of respiratory neuron to chemoreceptors afferents may then be considered in the context of this direct interaction as well as the network interactions of the various cells.