D. W. Richter

Synaptic mechanisms involved in the inspiratory modulation of vagal cardio‐inhibitory neurones in the cat.

The respiratory modulation of the activity of vagal cardio‐inhibitory neurones of the nucleus ambiguus of the cat has been investigated by electrophysiological and neuropharmacological techniques. All twenty‐four vagal efferent neurones studied had axons with conduction velocities indicative of B fibres and projected to the right cardiac branches of the vagus. Their spontaneous or DL‐homocysteic acid (DLH)‐evoked activity showed a marked reduction during the phase of inspiration and all showed signs of receiving a baroreceptor input. Ionophoretic application of DLH always excited cardiac vagal motoneurones (c.v.m.s). Application of acetylcholine to these same cells provoked a decrease in firing rate in twelve of the fifteen neurones tested. In ten of these twelve cells simultaneous application of atropine antagonized the effect of acetylcholine. Atropine applied alone enhanced neuronal firing, particularly in inspiration. Stable intracellular recordings have been made from two c.v.m.s. These were inhibited during inspiration. Input resistance fell markedly during inspiration and injection of chloride reversed this wave of hyperpolarization to a wave of depolarization indicating that this resulted from chloride‐mediated inhibitory post‐synaptic potentials (i.p.s.p.s). These c.v.m.s were activated during Stage I expiration, and showed a weak and variable wave of inhibition in Stage II expiration. Pulse‐rhythmic depolarizing potentials were reduced in their amplitudes during the periods of decreased neurone input resistance. It is concluded that c.v.m.s receive an excitatory input during post‐inspiration and a powerful inhibitory synaptic input during inspiration. The implications of these observations for the physiology of cardiorespiratory reflexes are discussed.

Reflex prolongation of stage I of expiration

Experiments were performed on anesthetized cats to test the theory that the interval between phrenic bursts is comprised of two phases, stage I and stage II of expiration. Evidence that these represent two separate neural phases of the central respiratory rhythm was provided by the extent to which stage duration is controlled individually when tested by superior laryngeal, vagus and carotid sinus nerve stimulation. Membrane potential trajectories of bulbar postinspiratory neurons were used to identify the timing of respiratory phases.Stimulation of the superior laryngeal, vagus and carotid sinus nerves during stage I of expiration prolonged the period of depolarization in postinspiratory neurons without significantly changing the durations of either stage II expiratory or inspiratory inhibition, indicating a fairly selective prolongation of the first stage of expiration. Changes in subglottic pressure, insufflation of smoke into the upper airway, application of water to the larynx or rapid inflation of the lungs produced similar effects. Sustained tetanic stimulation of superior laryngeal and vagus nerves arrested the respiratory rhythm in stage I of expiration. Membrane potentials in postinspiratory, inspiratory and expiratory neurons were indicative of a prolonged postinspiratory period. Thus, such an arrhythmia can be described as a postinspiratory apneic state of the central oscillator. The effects of carotid sinus nerve stimulation reversed when the stimulus was applied during stage II expiration. This was accompanied by corresponding changes in the membrane potential trajectories in postinspiratory neurons.The results manifest a ternary central respiratory cycle with two individually controlled phases occurring between inspiratory bursts.

Studies on the synaptic interconnection between bulbar respiratory neurones of cats.

In cats anaesthetized with pentobarbital, medullary respiratory neurones of both dorsal and ventral populations were recorded intracellularly with 1 mol·l−1 KCl-electrodes. The neurones were classified according to the projection of their axons to the spinal cord (bulbospinal neurones) or to the vagal nerves (vagal neurones). Those neurones which could not be activated antidromically (NAA-neurones) by either procedure were subdivided into (inspiratory) Rβ-neurones, which were monosynaptically excited by lung stretch receptor afferents, and into inspiratory and expiratory NAA-neurones, which did not receive a direct synaptic input, from these afferents.All types of neurone investigated revealed postsynaptic activity during both inspiration and expiration. The periods when synaptic activity was minimal were the periods of transition between respiratory phases.The input resistance of most respiratory neurones varied in parallel with the respiratory cycle. A drastic fall of the input resistance during expiration was observed in Rβ-neurones and in some inspiratory vagal neurones. This was not seen in inspiratory bulbospinal neurones.In stable intracellular recordings, periodic postsynaptic inhibition was demonstrated in 52 of 53 respiratory neurones by IPSP reversal following chloride injection. Maximal membrane potential then was generally reached during one of the periods of respiratory phase transition. Reasons for the failure of others to demonstrate these IPSPs are presented and discrepancies between other findings and these are discussed. It is concluded that reciprocal inhibition between bulbar respiratory neurones does exist and is a general phenomenon.It is argued that reciprocal inhibition is the fundamental mechanism underlying respiratory gating of afferent inputs.The probable existence of recurrent inhibition is inferred from the changes in the pattern of membrane depolarization during the active period of neurones.

The differential organization of medullary post-inspiratory activities

Membrane potential trajectories of 68 bulbar respiratory neurones from the peri-solitary and peri-ambigual areas of the brain-stem were recorded in anaesthetized cats to explore the synaptic influences of post-inspiratory neurones upon the medullary inspiratory network.A declining wave of inhibitory postsynaptic potentials resembling the discharge of postpinspiratory neurones was seen in both bulbospinal and non-bulbospinal inspiratory neurones, including alpha- and beta-inspiratory, early-inspiratory, late-inspiratory and ramp-inspiratory neurones.Activation of laryngeal and high-threshold pulmonary receptor afferents excited bulbar post-inspiratory neurones, whilst in the case of inspiratory neurones such stimulation produced enhanced postsynaptic inhibition during the same period of the cycle. Activation of post-inspiratory neurones and enhanced post-inspiratory inhibition of inspiratory bulbospinal neurones was accompanied by supression of the after-discharge of phrenic motoneurones.These results suggest that a population of post-inspiratory neurones exerts a widespread inhibitory function at the lower brain-stem level. Implications of such an inhibitory function for the organization of the respiratory network are discussed in relation to the generation of the respiratory rhythm.

Morphological and electrical description of medullary respiratory neurons of the cat

SummaryThe activity of 32 respiratory neurons located within the retroambigual region of the lower brain stem was recorded intracellulary and the neurons were stained using Procion Yellow. Inspiratory and expiratory bulbospinal (BS), vagal (V) neurons were identified by stimulation within the ventrolateral portions of the cervical spinal cord and the vagal nerves. Another group of inspiratory and expiratory neurons was not antidromically activated by such stimulations (NAA-neurons).A general feature of the retroambigual region seemed to be that neurons were clustered with rich overlapping of the dendritic trees. Dendritic overlapping was also seen at neighbouring respiratory neurons which had been stained by Procion Yellow. These clusters of cells were surrounded by a mass of myelinated axons, which ran predominantly in a longitudinal direction.The morphology of the cell somata, axons, and the dendritic trees was reconstructed from histological sections. The shapes of individual respiratory neurons of the different types were then investigated. The mean surface area of the somata was 5900 μm2 in V-neurons, 3800 μm2 in BS-neurons and 1800 μm2 in NAA-neurons. The dendritic tree extended over a length of up to 400 μm in BS- and V-neurons and over a length of up to 200 μm in NAA-neurons. The dendrites of respiratory neurons spread mainly in dorsomedial and ventrolateral directions. The axons of BS- and V-neurons coursed dorsomedially and acquired a myelin sheath some 30–50 μm away from the cell body.The input resistance of the neurons and the time constants for passive voltage decay transients produced by current pulse injection were estimated during the different periods of the respiratory cycle. The length and diameter of the dendritic segments were measured and the combined dendritic trunk parameter (Σd3/2) was calculated. This parameter decreased with increasing electrotonic length of the dendrites.These results, in combination with the morphological data, indicate that respiratory neurons receive a tonic synaptic input. Additionally, these measurements have revealed cyclic changes of synaptic input to remote portions of the dendritic tree which seem to exert a significant influence on neuronal activity.

Voltage-dependent currents in neurones of the nuclei of the solitary tract of rat brainstem slices

AbstractNeurones within the ventral and ventrolateral nuclei of the solitary tract were analyzed under single-electrode current- and voltage-clamp conditions in rat brainstem slices. We present direct and indirect evidence for the existence of five different sorts of membrane currents:1.a tetrodotoxin-sensitive sodium current,2.a tetrodotoxin-resistant calcium current,3.a calcium-dependent potassium current,4.a non-inactivating potassium current which is inhibited by muscarine,5.an inactivating potassium current, which is inhibited by 4-aminopyridine. These membrane properties do not produce spontaneous bursting in these neurones. Assuming that neurones with such properties belong to the respiratory network, we discuss how conductances of this type may be involved in mechanisms regulating central respiratory activity.

Changes in extracellular potassium during the spontaneous activity of medullary respiratory neurones

In 34 cats, the changes in extracellular potassium ion activity (aK) and extracellular spike activity within the pool of respiratory neurones in the dorsormedial and ventrolateral medulla were recorded using microelectrodes filled with a liquid potassium ion exchange resin. Cyclic changes in aK which parallel central respiratory activity were restricted to those regions where respiratory neurones are known to be localized. The largest changes in aK (0.1–0.3 mmol · l−1) were found within the ventral pool of inspiratory neurones. The aK increased during inspiration in parallel with the pattern of phrenic nerve activity. The smallest changes in aK (0.02–0.06 mmol · l−1) were observed within the ventral pool of expiratory neurones. Here, aK showed a transient increase during both inspiration and expiration. Within the dorsal pool of inspiratory neurones, small fluctuations of aK were observed paralleling phrenic nerve activity and the afferent discharge of the intact vagal nerves. After the vagal nerves were cut, the changes in aK then paralleled phrenic nerve activity. The variations in aK within the ventral pool of respiratory neurones did not change after bilateral section of vagal nerves.Repetitive stimulation of the vagal nerves (0.1–0.5V, 0.05 ms) produced an increase in aK only within the dorsal pool of inspiratory neurones, whereas repetitive spinal cord stimulation (5–10V, 0.05 ms) resulted in an increase of aK within the ventral pool of respiratory neurones.The amplitude of the cyclic changes in aK increased significantly whenever the electrode approached individual respiratory neurones as verified by the amplitude and shape of the spikes recorded by the reference barrel. The maximal changes in aK then reached a peak amplitude of 1.3–1.5 mmol · l−1, the pattern of aK changes resembling that measured within the pools of neurones.The aK started to rise prior to the discharge of action potentials, indicating that the efflux of K+-ions was produced as a consequence of synaptic transmission. The functional importance of these changes in extracellular potassium is discussed.

Evidence for a monosynaptic connection between slowly adapting pulmonary stretch receptor afferents and inspiratory beta neurones

The synaptic connection between slowly adapting pulmonary stretch receptor afferents and inspiratory neurones within a region ventral to the tractus solitarius was determined using intracellular recording and spike triggered averaging techniques.When the vagus nerve was stimulated at intensities eliciting a Hering-Breuer reflex, the difference in mean latency between centrally recorded action potentials of slowly adapting pulmonary stretch receptor afferents and e.p.s.p.s of inspiratory beta neurones was 0.2 ms. This difference is indicative of a monosynaptic connection.Extracellular single unit spikes of slowly adapting pulmonary stretch receptors recorded from the nodose ganglion were used to trigger the averaging of synaptic noise recorded from inspiratory neurones. A prominent wave of synaptic depolarization was observed in all inspiratory beta neurones even when a small number of sweeps were averaged. This depolarization was absent from inspiratory alpha neurones.The shape indices of these depolarizations are consistent with a monosynaptic connection between slowly adapting pulmonary stretch receptor afferents and inspiratory beta neurones. In addition, the data raise the possibility that this connection is multiple and distributed.

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 μl 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.

Presynaptic depolarization in myelinated vagal afferent fibres terminating in the nucleus of the tractus solitarius in the cat

The presynaptic influences that act on terminals of slowly adapting lung stretch receptor afferents and aortic baroreceptor afferents within the nucleus of the solitary tract were assessed using intracellular recording and antidromic stimulation techniques.Central respiratory influences on the axcitability of lung stretch receptor terminals were observed in 29% (4 of 14) of measurements. These were confirmed in intracellular recordings where membrane depolarizations in synchrony with phrenic nerve discharge were seen in 17% (4 of 24) of fibres. In three cases membrane depolarization also occurred synchronously with artificial lung inflation.Neither tests of excitability nor intracellular recording revealed any evidence for equivalent presynaptic influences on 16 myelinated aortic baroreceptor terminals.Stimulation of the superior laryngeal nerve evoked depolarizations in 50% (7 of 14) of lung stretch receptor terminals. These took the form of complex waves of depolarization with both short (3–8 ms) and long latency (27–35 ms) components. The amplitude of the long latency response increased during the period of phrenic nerve discharge, i.e. during “central inspiration”.These effects are discussed in relation to the central respiratory influences on both respiratory and cardiovascular reflexes.