Paraventricular Nucleus Modulates Excitatory Cardiovascular Reflexes during Electroacupuncture.

The paraventricular nucleus (PVN) regulates sympathetic outflow and blood pressure. Somatic afferent stimulation activates neurons in the hypothalamic PVN. Parvocellular PVN neurons project to sympathoexcitatory cardiovascular regions of the rostral ventrolateral medulla (rVLM). Electroacupuncture (EA) stimulates the median nerve (P5-P6) to modulate sympathoexcitatory responses. We hypothesized that the PVN and its projections to the rVLM participate in the EA-modulation of sympathoexcitatory cardiovascular responses. Cats were anesthetized and ventilated. Heart rate and mean blood pressure were monitored. Application of bradykinin every 10-min on the gallbladder induced consistent pressor reflex responses. Thirty-min of bilateral EA stimulation at acupoints P5-P6 reduced the pressor responses for at least 60-min. Inhibition of the PVN with naloxone reversed the EA-inhibition. Responses of cardiovascular barosensitive rVLM neurons evoked by splanchnic nerve stimulation were reduced by EA and then restored with opioid receptor blockade in the PVN. EA at P5-P6 decreased splanchnic evoked activity of cardiovascular barosensitive PVN neurons that also project directly to the rVLM. PVN neurons labeled with retrograde tracer from rVLM were co-labeled with μ-opioid receptors and juxtaposed to endorphinergic fibers. Thus, the PVN and its projection to rVLM are important in processing acupuncture modulation of elevated blood pressure responses through a PVN opioid mechanism

Studies on the respiratory modulation of sympathetic activity

Sympathetic activity is modulated by central respiratory drive. Studies using whole nerve recordings in the rat have demonstrated different patterns of respiratory modulation in various sympathetic nerves. These regional differences in the discharge patterns of sympathetic outflows may result from either varying proportions of sympathetic neurones with a particular respiratory-related discharge pattern contributing to each whole-nerve activity or sympathetic preganglionic neurones (SPNs) projecting into different nerves having characteristic respiratory modulations. The present study has investigated the respiratory-related discharge patterns of a group of SPNs projecting to the lumbar sympathetic chain (LSC). Furthermore, the hypothesis that caudal raphe nuclei (raphe obscurus, pallidus and magnus) convey central respiratory drive onto sympathetic outflow has been examined. In anaesthetized and vagotomized rats extracellular recordings were made from identified SPNs projecting to or through the lumbar sympathetic chain between L4 and L5 ganglia, and from caudal raphe neurones with axons projecting to the spinal cord. The respiratory-related firing patterns were analysed. Differences in patterns of respiratory modulation and the proportion of SPNs with a certain pattern of respiratory modulation were found between SPNs recorded in the present study and SPNs located in upper thoracic spinal segments reported previously. These findings provide an explanation of the regional differences of respiratory modulation in various sympathetic nerves. Many caudal raphe-spinal neurones with respiratory-related activity could be activated antidromically from the area of the intermediolateral cell column (IML) and activity in some of these neurons correlated to the 2 to 6 Hz rhythm of cervical sympathetic activity. The findings are consistent with the idea that caudal raphe neurones within the region from which I recorded in this study are part of a supraspinal network that contributes to the 2 to 6 Hz component of sympathetic nerve activity. Therefore some raphe-spinal neurones may relay both "respiratory" and "sympathetic" rhythmic components to the sympathetic outflow. These spinally- projecting neurones in caudal raphe nuclei are different from those in the rostral ventrolateral medulla (RVLM) as they have no baroreceptor-related activity. Additionally, they do not have the "typical" characteristics of 5-HT containing neurones which have slow conduction velocities, and slow regular firing characteristics: the majority had small myelinated axons as indicated by their conduction velocities and relatively high discharge rates and irregular firing characteristics

Antinociceptive actions of descending catecholaminergic tracts on dorsal horn somatosensory neurones

Ionophoretically-applied dopamine and noradrenaline selectively inhibited the nociceptive responses of multireceptive somatosensory dorsal horn neurones, whilst non-nociceptive responses, spontaneous activity and activity evoked by an ionophoretically-applied excitatory amino acid, DL-homocysteic acid were unaffected. Many of the neurones tested had long ascending projections, capable of transmitting nociceptive information to supraspinal sites; in the rat (spinothalamic tract neurones) and in the cat (spinocervical tract neurones).The use of ionophoretically-applied receptor-specific agonists and antagonists demonstrated that the actions of noradrenaline and dopamine were pharmacologically distinct. The selective antinociceptive action produced by noradrenaline was mediated by an 06,-adrenoreceptor, whilst the selective antinociceptive effect of dopamine was mediated by a dopamine receptor.A glyoxylic-acid histofluorescence study was undertaken to ascertain the optimal stereotaxic placement of stimulating electrodes, in the regions of those dopamine cell groups (A9 and All) that have been considered to provide a spinal projection.Focal electrical stimulation in the region of the All dopamine cell group evoked a selective antinociceptive effect on multireceptive dorsal horn neurones in the rat. This stimulus-evoked effect was rapidly and consistently reversed by ionophoresis of the D₂ dopamine-receptor antagonist, sulpiride, in the vicinity of the dorsal horn neurone being tested, whilst an opiate antagonist (naloxone) and an α₂-antagonist (RX781094) had no effect. Using the same parameters, focal electrical stimulation in the region of the A9 dopamine cell group did not affect the evoked responses of any multireceptive neurones tested.The results of this study present data supporting selective antinociceptive roles for dopamine and noradrenaline at the spinal level. The All dopamine cell group was demonstrated as a supraspinal source of a selective antinociceptive effect, mediated by dopamine at the level of the dorsal horn

The origins of electrical discharge patterns in the main olfactory bulb of the rat

Landmark discoveries made in the olfactory bulb have formed the basis of much of our understanding of other brain regions. In fact, it was in the olfactory bulb that the first example of a dendrodentritic synapse in the mammalian central nervous system was found. The olfactory bulb is rich in a diverse selection of neurotransmitters, and with the bonus that the bulb is relatively easy to access, it provides an excellent model in which to study neural networks.The aim of this PhD project was to study the neural pathway that is thought to connect the olfactory bulb to the supraoptic nucleus of the hypothalamus. To understand the input to the supraoptic nucleus from the olfactory system we sought to determine the discharge frequency and firing pattern of the output neurones, mitral cells. The electrical activity of single neurones was recorded extracellularly from the olfactory bulb of anaesthetised rats. Mitral cells were identified electrophysiologically by antidromic activation following stimulation of the lateral olfactory tract and observation of bulbar field potentials. It became apparent that the mitral cells consistently showed a spontaneous patterned discharge that has not been previously reported and we have described this pattern in terms of three separate levels of bursting behaviour.What we have termed the 'gross' phasic pattern was displayed by all mitral cells and consisted of a characteristic slow, cyclic firing pattern with peaks of activity occurring with a constant periodicity, burst lengths lasted for approximately two minutes with equal periods of quiescence separating the bursts. Some mitral cells (53 %) show distinct silent periods between bursts of high activity others (47 %) simply show a reduced rate of activity between bursts. Auto -correlation plots show that within this overall phasic pattern is a respiratory driven bursting activity, the activity of mitral cells increases during the inspiratory phase as air is drawn over the olfactory receptors in the nasal mucosa. Plotting the instantaneous frequency of mitral cell activity reveals the third bursting pattern, exhibited by 57% of mitral cells recorded. This shows that during each long burst of activity the mitral cell fires at two distinct frequencies, the lower frequency is in the range 0 -50Hz and the high frequency firing is in the range 100- 250Hz. In 84% of the bursts that showed two distinct firing frequencies there was a delay in the onset of the higher frequency mode, at the start of each peak of activity. Mitral cells have been shown to be capable of initiating and propagating action potentials from their distal dendrites, as well as from the conventional initiation site at the soma -axon hillock region. It is proposed in this thesis that the high frequency firing mode described might be generated in the mitral cell dendrites.The mitral cell is involved in complex interactions with both neighbouring mitral cells and granule cells that provide for lateral and reciprocal inhibition respectively. Granule cells are the most numerous of the various types of interneurone in the bulb and their firing pattern was found to be non-phasic and at only one frequency mode. Following stimulation of the lateral olfactory tract mitral cells exhibited a period of inhibition following the stimulus pulse. This is consistent with the general consensus that upon activation, mitral cells activate granule cells, which in turn feedback to inhibit the mitral cells (reciprocal inhibition). Extracellular recordings of mitral cell activity were also made in a slice preparation of the olfactory bulb. It was discovered that in vitro the mitral cells did not discharge in a slow, phasic pattern and the high frequency firing seen in vivo was not evident. During the slice preparation many of the long lateral dendrites of the mitral cells are unavoidably removed and this may disturb the local interactions and thereby alter the discharge pattern.Once the discharge pattern of olfactory neurones was determined these parameters were then used as a basis for the stimulation of the lateral olfactory tract and the effect on supraoptic neurone activity determined by studying the distribution of Fos -positive cells. Two stimulation protocols were used both were strong stimuli applied unilaterally, in different formats. The first was a short burst at a high frequency to mimic an acute, strong output from the olfactory bulb and the second Abstract x was a prolonged stimulation used to disrupt the output discharge pattern. The literature suggests that the connection between the olfactory bulb and the supraoptic nucleus is unilateral, monosynaptic and terminates in the ventro -lateral dendritic region of the supraoptic nucleus. Following prolonged stimulation of the lateral olfactory tract there was a significant increase (p <0.01) in Fos expression in the supraoptic nucleus on both the ipsilateral and contralateral sides which suggests that the pathway between the olfactory bulb and the supraoptic nucleus may be more complex than initially thought. Areas of the brain known to receive strong olfactory input, such as the piriform cortex, showed a unilateral increase in Fos expression following the brief pulse of stimulation.Administration of morphine during parturition interrupts the progress of parturition by inhibiting oxytocin release. The olfactory bulb is highly active at the time of parturition and shows dense expression of mu- and kappa opioid receptors, and so the possibility that morphine may impair oxytocin release in part by blocking the input from the olfactory bulb was considered. The effect of morphine and its antagonist, naloxone on the discharge pattern of mitral cells was studied in both the in vivo and in vitro preparations. In vivo morphine was seen to have a subtle effect in that it inhibited the high frequency firing but did not significantly alter the overall firing rate or periodicity of bursts, this effect was irreversible. However, in vitro morphine fully inhibited mitral cell activity which returned to pre -morphine rates following the administration of naloxone. The discrepancy between the two sets of data may be a dose issue. In vivo the drugs were administered via an intravenous route which may have led to a reduced concentration of the drug evoking a response from the mitral cells compared to the concentration of the drug that the mitral cells in vitro were exposed to. It may also be due to the reduced local circuitary of the mitral cell in the in vitro preparation, causing the mitral cell to become more susceptible to the effects of morphine

Phasic Firing in Vasopressin Cells: Understanding Its Functional Significance through Computational Models

Vasopressin neurons, responding to input generated by osmotic pressure, use an intrinsic mechanism to shift from slow irregular firing to a distinct phasic pattern, consisting of long bursts and silences lasting tens of seconds. With increased input, bursts lengthen, eventually shifting to continuous firing. The phasic activity remains asynchronous across the cells and is not reflected in the population output signal. Here we have used a computational vasopressin neuron model to investigate the functional significance of the phasic firing pattern. We generated a concise model of the synaptic input driven spike firing mechanism that gives a close quantitative match to vasopressin neuron spike activity recorded in vivo, tested against endogenous activity and experimental interventions. The integrate-and-fire based model provides a simple physiological explanation of the phasic firing mechanism involving an activity-dependent slow depolarising afterpotential (DAP) generated by a calcium-inactivated potassium leak current. This is modulated by the slower, opposing, action of activity-dependent dendritic dynorphin release, which inactivates the DAP, the opposing effects generating successive periods of bursting and silence. Model cells are not spontaneously active, but fire when perturbed by random perturbations mimicking synaptic input. We constructed one population of such phasic neurons, and another population of similar cells but which lacked the ability to fire phasically. We then studied how these two populations differed in the way that they encoded changes in afferent inputs. By comparison with the non-phasic population, the phasic population responds linearly to increases in tonic synaptic input. Non-phasic cells respond to transient elevations in synaptic input in a way that strongly depends on background activity levels, phasic cells in a way that is independent of background levels, and show a similar strong linearization of the response. These findings show large differences in information coding between the populations, and apparent functional advantages of asynchronous phasic firing

Cutaneous C-fibres in the rat and the rabbit - How efferent actions and axonal properties vary with functional class

The aim of this study was to investigate further some efferent actions and axonal properties of the unmyelinated fibres innervating rabbit and rat skin. This investigation can be separated into two parts. Firstly, single unit studies were carried out to determine which functional class(es) or sub-class(es) of C-fibre are responsible for antidromic vasodilatation in both rabbit and rat skin. The findings of these single unit studies were compared with the flare responses of the skin to noxious mechanical and heat stimuli. Secondly, activity-dependent slowing of conduction velocity and axonal spike shape were examined in identified cutaneous C-fibres in the rat in order to determine whether such axonal properties could be used to identify the different functional classes of C-fibre. For the antidromic vasodilatation study, fine filaments were dissected from the cut proximal end of the saphenous nerve in anaesthetized rabbits and rats. Individual C-fibres (conduction velocity <2m/s) were classified into functional groups according to their responses to mechanical and thermal stimulation. The threshold for electrical stimulation of individual C-fibres was determined using the collision technique. Filaments were antidromically electrically stimulated at intensities sufficient to excite the conducting C-fibres, and skin blood flow was monitored before, during and after filament stimulation using laser Doppler perfusion imaging or laser Doppler flowmetry. In both species, the only C-fibres capable of producing a detectable vasodilator response following antidromic stimulation were nociceptive in nature, and in all cases the area of vasodilatation coincided well with the afferent receptive field. However, not all nociceptors produced a detectable vasodilatation, and it seems that a sub-group of polymodal and heat nociceptors are responsible for the efferent action of antidromic vasodilatation in rabbit and rat skin. Flare responses in rabbit and rat skin were only detected following mechanical and heat stimuli within noxious ranges. The spread of the flare responses, together with the sizes of the afferent and efferent receptive fields of individual C-units, provide support for the axon reflex mechanism for the production of flare and antidromic vasodilatation. The percentage slowing of conduction velocity was calculated following a standard electrical tetanus in identified C-fibres dissected from the saphenous nerve in anaesthetized rats. Nociceptive C-fibres showed a greater slowing of conduction velocity than non-nociceptive fibres, and moreover, one could separate the two classes of non-nociceptive afferent C-fibres found in the rat saphenous nerve (the mechanoreceptors and cold thermoreceptors) on the basis of their conduction velocity slowing. In addition, activity-dependent slowing of conduction velocity could be used to differentiate between the afferent and non-afferent populations of inexcitable C-fibres. Spike shapes of functionally classified C-fibres were recorded extracellularly using standardized filter settings, and some variations in spike shape in relation to receptor type were found. Polymodal nociceptors displayed wider spikes than mechanoreceptors, and cold thermoreceptor units tended to have monophasic spikes. Also, the spontaneously active sympathetic efferent C-fibres tended to have spikes of relatively long duration. The use of axonal properties such as activity-dependent slowing of conduction velocity and spike shape to differentiate nociceptors from non-nociceptors has great potential in experiments where axons are isolated from their terminals

Cellular specialization of synaptic integration in a mammalian sympathetic ganglion

Sympathetic ganglia are widely viewed as simply relays that are essential to convey neural activity from spinal preganglionic neurons to distinct peripheral targets. However, recent studies indicate that synaptic integration in sympathetic ganglia is more complex than that of a simple excitatory relay. It is proposed that synaptic organization of each functional subset of sympathetic ganglion cells is specialized to generate a unique synaptic gain function, thereby allowing for differential control of specific target modalities. This dissertation describes cellular specialization of some critical determinants of synaptic gain in rat superior cervical ganglion (SCG) neurons. The work was first focused on identifying presynaptic stimulus threshold and NPY immunoreactivity as neuronal classification criteria of secretomotor, pilomotor and vasoconstrictor cells. The results here show that these three functional phenotypes of neurons are indistinguishable in terms of synaptic convergence. Furthermore, norepinephrine (NE) causes different modulatory effects upon pre and postsynaptic ¦Á2-adrenergic receptors in these cell types. Collectively, this work characterizes cellular specialization of synaptic convergence and NE neuromodulatory mechanism that are involved in synaptic integration in the rat SCG

Actions and interactions of high pressure and general anaesthetics in rat hippocampal Ca1 neurones

The thesis is divided into two experimental sections: the first concerns the actions of general anaesthetics on rat hippocampal CA1 neurones at atmospheric pressure. The second investigates some properties of CA1 neurones under high helium pressure. The general anaesthetics studied at one atmosphere were enflurane, isoflurane, halothane, ketamine and methohexitone. Antidromic field potential measurements were taken in the absence and presence of the anaesthetics in order to assess changes in axonal/somatic excitability. Accommodation behaviour of CA1 neurones was also investigated in intracellular experiments with the above anaesthetics. The principal findings were that the anaesthetics studied decreased the amplitude of the antidromic field potential and induced hyperpolarization, with the exception of ketamine which enhanced antidromic responses at low concentration and had mixed effects upon the resting potential. Halothane also induced a second antidromic population spike. The inhalation anaesthetics (enflurane, isoflurane, halothane) all blocked accommodation. Ketamine was found to slightly compromise accommodation, whilst methohexitone had mixed effects. A voltage-clamp study indicated that enflurane reduced the M-current of CA1 neurones. CA1 neurone responses to helium pressure (up to 13.3MPa) were investigated using a purpose built pressure chamber designed to facilitate intracellular recording. In field potential experiments antidromic and orthodromic responses (to both single and paired pulses) were studied at one atmosphere and following compression. Responses were found to be mixed at elevated pressure. Some preparations were found to be unaffected by pressure whilst others became more excitable. Ketamine and methohexitone were found to have similar actions at 10MPa to their actions at 0.1 MPa. Intracellular measurements were made at pressure (5MPa and 10MPa). Resting membrane potential, input resistance and threshold potential were found to be unaffected by pressure. High pressure was found to block accommodation and reduce the associated AHP in CA1 neurones

Signal Processing and Distribution in Cortical?Brainstem Pathways for Smooth Pursuit Eye Movements

Smooth pursuit (SP) eye movements are used to maintain the image of a moving object relatively stable on the fovea. Even when tracking a single target over a dark background, multiple areas including frontal eye fields (FEF) and middle temporal (MT) and medial superior temporal (MST) cortex contribute to converting visual signals into initial commands for SP. Signals in the cortical pursuit system reach the oculomotor cerebellum through brainstem centers including the dorsolateral pontine nucleus (DLPN), nucleus reticularis tegmenti pontis (NRTP), and pretectal nucleus of the optic tract (NOT). The relative information carried in these parallel pathways remains to be fully defined. We used multiple linear?regression modeling to estimate the relative sensitivities of cortical (MST, FEF), pontine (NRTP, DLPN), and NOT neurons to eye? and retinal?error parameters (position, velocity, and acceleration) during step?ramp SP of macaques (Macaca mulatta). We found that a large proportion of pursuit?related MST and DLPN neurons were most sensitive to eye?velocity or retinal error velocity. In contrast, a large proportion of FEF and rostral NRTP neurons were most sensitive to eye acceleration. Visual neurons in MST, DLPN, and NOT were most sensitive to retinal image velocity