Preganglionic Inputs Research Articles

Secondhand Smoke Decreased Excitability and Altered Action Potential Characteristics of Cardiac Vagal Neurons in Mice.

Background: Secondhand smoke (SHS), a major indoor pollutant, is a significant risk factor for cardiovascular morbidity and mortality including arrhythmias and sudden cardiac death. Exposure to SHS can produce autonomic imbalance, as evidenced by reduced heart rate variability (HRV)—a clinical metric of cardiac vagal regulation. Currently, the mechanisms through which SHS changes the vagal preganglionic neuronal inputs to the heart to produce this remains unknown.Objectives: To characterize the effect of SHS on both the excitability and action potential (AP) characteristics of anatomically identified cardiac vagal neurons (CVNs) in the nucleus ambiguus and examine whether SHS alters small conductance calcium-activated potassium (SK) channel activity of these CVNs.Methods: Adult male mice were exposed to four weeks of filtered air or SHS (3 mg/m3) 6 h/day, 5 day/week. Using patch-clamp recordings on identified CVNs in brainstem slices, we determined neuronal excitability and AP characteristics with depolarizing step- and ramp-current injections.Results: Four weeks of SHS exposure reduced spiking responses to depolarizing current injections and increased AP voltage threshold in CVNs. Perfusion with apamin (20 nM) magnified these SHS-induced effects, suggesting reduced SK channel activity may serve to minimize the SHS-induced decreases in CVNs excitability. Medium afterhyperpolarization (a measurement of SK channel activity) was smaller in the SHS group, further supporting a lower SK channel activity. AP amplitude, rise rate, fast afterhyperpolarization amplitude (a measurement of voltage-gated channel activity), and decay rate were higher in the SHS group at membrane voltages more positive to 0 mV, suggesting altered inactivation properties of voltage-dependent channels underlying APs.Discussion: SHS exposure reduced neuronal excitability of CVNs with compensatory attenuation of SK channel activity and altered AP characteristics. Neuroplasticity of CVNs could blunt regulatory cardiac vagal signaling and contribute to the cardiovascular consequences associated with SHS exposure, including reduced HRV.

Spatiotemporal mapping of sensory and motor innervation of the embryonic and postnatal mouse urinary bladder

The primary function of the urinary bladder is to store urine (continence) until a suitable time for voiding (micturition). These distinct processes are determined by the coordinated activation of sensory and motor components of the nervous system, which matures to enable voluntary control at the time of weaning. Our aim was to define the development and maturation of the nerve-organ interface of the mouse urinary bladder by mapping the organ and tissue distribution of major classes of autonomic (motor) and sensory axons. Innervation of the bladder was evident from E13 and progressed dorsoventrally. Increasing defasciculation of axon bundles to single axons within the muscle occurred through the prenatal period, and in several classes of axons underwent further maturation until P7. Urothelial innervation occurred more slowly than muscle innervation and showed a clear regional difference, from E18 the bladder neck having the highest density of urothelial nerves. These features of innervation were similar in males and females but varied in timing and tissue density between different axon classes. We also analysed the pelvic ganglion, the major source of motor axons that innervate the lower urinary tract and other pelvic organs. Cholinergic, nitrergic (subset of cholinergic) and noradrenergic neuronal cell bodies were present prior to visualization of these axon classes within the bladder. Examination of cholinergic structures within the pelvic ganglion indicated that connections from spinal preganglionic neurons to pelvic ganglion neurons were already present by E12, a time at which these autonomic ganglion neurons had not yet innervated the bladder. These putative preganglionic inputs increased in density prior to birth as axon terminal fields continued to expand within the bladder tissues. Our studies also revealed in numerous pelvic ganglion neurons an unexpected transient expression of calcitonin gene-related peptide, a peptide commonly used to visualise the peptidergic class of visceral sensory axons. Together, our outcomes enhance our understanding of neural regulatory elements in the lower urinary tract during development and provide a foundation for studies of plasticity and regenerative capacity in the adult system.

Role of the superior salivatory nucleus in parasympathetic control of choroidal blood flow and in maintenance of retinal health

The vasodilatory pterygopalatine ganglion (PPG) innervation of the choroid is under the control of preganglionic input from the superior salivatory nucleus (SSN), the parasympathetic portion of the facial motor nucleus. We sought to confirm that choroidal SSN drives a choroid-wide vasodilation and determine if such control is important for retinal health. To the former end, we found, using transscleral laser Doppler flowmetry, that electrical activation of choroidal SSN significantly increased choroidal blood flow (ChBF), at a variety of choroidal sites that included more posterior as well as more anterior ones. We further found that the increases in ChBF were significantly reduced by inhibition of neuronal nitric oxide synthase (nNOS), thus implicating nitrergic PPG terminals in the SSN-elicited ChBF increases. To evaluate the role of parasympathetic control of ChBF in maintaining retinal health, some rats received unilateral lesions of SSN, and were evaluated functionally and histologically. In eyes ipsilateral to choroidal SSN destruction, we found that the flash-evoked scotopic electroretinogram a-wave and b-wave peak amplitudes were both significantly reduced by 10 weeks post lesion. Choroidal baroregulation was evaluated in some of these rats, and found to be impaired in the low systemic arterial blood pressure (ABP) range where vasodilation normally serves to maintain stable ChBF. In retina ipsilateral to SSN destruction, the abundance of Müller cell processes immunolabeled for glial fibrillary acidic protein (GFAP) and GFAP message were significantly upregulated. Our studies indicate that the SSN-PPG circuit mediates parasympathetic vasodilation of choroid, which appears to contribute to ChBF baroregulation during low ABP. Our results further indicate that impairment in this adaptive mechanism results in retinal dysfunction and pathology within months of the ChBF disturbance, indicating its importance for retinal health.

Functional segregation within the pelvic nerve of male rats: a meso- and microscopic analysis.

The pelvic splanchnic nerves are essential for pelvic organ function and have been proposed as targets for neuromodulation. We have focused on the rodent homologue of these nerves, the pelvic nerves. Our goal was to define within the pelvic nerve the projections of organ‐specific sensory axons labelled by microinjection of neural tracer (cholera toxin, subunit B) into the bladder, urethra or rectum. We also examined the location of peptidergic sensory axons within the pelvic nerves to determine whether they aggregated separately from sacral preganglionic and paravertebral sympathetic postganglionic axons travelling in the same nerve. To address these aims, microscopy was performed on the major pelvic ganglion (MPG) with attached pelvic nerves, microdissected from young adult male Sprague–Dawley rats (6–8 weeks old) and processed as whole mounts for fluorescence immunohistochemistry. The pelvic nerves were typically composed of five discrete fascicles. Each fascicle contained peptidergic sensory, cholinergic preganglionic and noradrenergic postganglionic axons. Sensory axons innervating the lower urinary tract (LUT) consistently projected in specific fascicles within the pelvic nerves, whereas sensory axons innervating the rectum projected in a complementary group of fascicles. These discrete aggregations of organ‐specific sensory projections could be followed along the full length of the pelvic nerves. From the junction of the pelvic nerve with the MPG, sensory axons immunoreactive for calcitonin gene‐related peptide (CGRP) showed several distinct patterns of projection: some projected directly to the cavernous nerve, others projected directly across the surface of the MPG to the accessory nerves and a third class entered the MPG, encircling specific cholinergic neurons projecting to the LUT. A subpopulation of preganglionic inputs to noradrenergic MPG neurons also showed CGRP immunoreactivity. Together, these studies reveal new molecular and structural features of the pelvic nerves and suggest functional targets of sensory nerves in the MPG. These anatomical data will facilitate the design of experimental bioengineering strategies to specifically modulate each axon class.

Physiological and pathophysiological firing properties of single postganglionic sympathetic neurons in humans.

It has long been known from microneurographic recordings in human subjects that the activity of postganglionic sympathetic axons occurs as spontaneous bursts, with muscle sympathetic nerve activity (MSNA) exhibiting strong cardiac rhythmicity via the baroreflex and skin sympathetic nerve activity showing much weaker cardiac modulation. Here we review the firing properties of single sympathetic neurons, obtained using highly selective microelectrodes. Individual vasoconstrictor neurons supplying muscle or skin, or sudomotor neurons supplying sweat glands, always discharge with a low firing probability (~30%) and at very low frequencies (~0.5 Hz). Moreover, they usually fire only once per cardiac interval but can fire greater than four times within a burst. Modeling has shown that this pattern can best be explained by individual neurons being driven by, on average, two preganglionic inputs. Unitary recordings of muscle vasoconstrictor neurons have been made in several pathophysiological states, including heart failure, hypertension, obstructive sleep apnea, bronchiectasis, chronic obstructive pulmonary disease, depression, and panic disorder. The augmented MSNA in each of these diseases features an increase in firing probability and discharge frequency of individual muscle vasoconstrictor neurons above that seen in healthy subjects, yet firing rates rarely exceed 1 Hz. However, unlike patients with heart failure, all patients with respiratory disease or panic disorder, and patients with hyperhidrosis, exhibited an increase in multiple within-burst firing, which emphasizes the different modes by which the sympathetic nervous system grades its output in pathophysiological states of high sympathetic nerve activity.

Ganglionic and non-ganglionic neurons in frog heart: do they differ?

This study was designed to compare the morphology of neurons in relation to their distance from the major nerve trunks in the heart of the frog Rana temporaria. Seventy-nine intracardiac neurons were labelled intracellularly with fluorescent markers Lucifer Yellow CH and Alexa Fluor 568. The neurons located on the extensions of the vagus nerve were considered as ganglionic, while neurons spread loosely at further distance from these extensions were considered as non-ganglionic. The mean area of the soma in ganglionic neurons was about 25% larger than in non-ganglionic neurons. Ganglionic neurons had a higher soma area/nucleus area ratio than non-ganglionic neurons. Although both the total number and the total length of dendrite-like processes was similar between the two groups, ganglionic neurons had significantly fewer dendrite-like processes from the soma (1.5±0.3 vs. 3.9±1.0; P<0.05) and shorter total length of these processes from the soma (63±18 μm vs. 178±51 μm; P<0.05). In conclusion, ganglionic and non-ganglionic frog intracardiac neurons exhibit substantial morphological differences. We hypothesize that these differences may indicate different projections or variations in the number of their preganglionic inputs.

The identification and neurochemical characterization of central neurons that target parasympathetic preganglionic neurons involved in the regulation of choroidal blood flow in the rat eye using pseudorabies virus, immunolabeling and conventional pathway tracing methods.

The choroidal blood vessels of the eye provide the main vascular support to the outer retina. These blood vessels are under parasympathetic vasodilatory control via input from the pterygopalatine ganglion (PPG), which in turn receives its preganglionic input from the superior salivatory nucleus (SSN) of the hindbrain. The present study characterized the central neurons projecting to the SSN neurons innervating choroidal PPG neurons, using pathway tracing and immunolabeling. In the initial set of studies, minute injections of the Bartha strain of the retrograde transneuronal tracer pseudorabies virus (PRV) were made into choroid in rats in which the superior cervical ganglia had been excised (to prevent labeling of sympathetic circuitry). Diverse neuronal populations beyond the choroidal part of ipsilateral SSN showed transneuronal labeling, which notably included the parvocellular part of the paraventricular nucleus of the hypothalamus (PVN), the periaqueductal gray, the raphe magnus (RaM), the B3 region of the pons, A5, the nucleus of the solitary tract (NTS), the rostral ventrolateral medulla (RVLM), and the intermediate reticular nucleus of the medulla. The PRV+ neurons were located in the parts of these cell groups that are responsive to systemic blood pressure signals and involved in systemic blood pressure regulation by the sympathetic nervous system. In a second set of studies using PRV labeling, conventional pathway tracing, and immunolabeling, we found that PVN neurons projecting to SSN tended to be oxytocinergic and glutamatergic, RaM neurons projecting to SSN were serotonergic, and NTS neurons projecting to SSN were glutamatergic. Our results suggest that blood pressure and volume signals that drive sympathetic constriction of the systemic vasculature may also drive parasympathetic vasodilation of the choroidal vasculature, and may thereby contribute to choroidal baroregulation during low blood pressure.

Sympathy for the ganglion.

Sympathy for the ganglion.

Segmental origins of cardiac sympathetic nerve activity in rats.

The segmental origins of cardiac sympathetic nerve activity (CSNA) were investigated in 8 urethane-anesthetized, artificially ventilated rats. The left upper thoracic sympathetic chain was exposed retropleurally after removing the heads of the second to fourth ribs. The preganglionic inputs to the chain from segments T1-T3 and the trunk distal to T3 were marked for later sectioning. CSNA was recorded conventionally, amplified, rectified and smoothed. Its mean level was quantified before and after each preganglionic input was cut, usually in rostro-caudal sequence. The level after all inputs were cut (i.e. noise and residual ECG pickup) was subtracted from previous measurements. The signal decrement from cutting each preganglionic input was then calculated as a percentage. CSNA in all rats depended on preganglionic drive from two or more segments, which were not always contiguous. Over the population, most preganglionic drive came from T3 and below, while the least came from T1. But there was striking inter-individual variation, such that the strongest drive to CSNA in any one rat could come from T1, T2, T3, or below T3. These findings provide new functional data on the segmental origins of CSNA in rats.

Inflammation in dorsal root ganglia after peripheral nerve injury: Effects of the sympathetic innervation

Following a peripheral nerve injury, a sterile inflammation develops in sympathetic and dorsal root ganglia (DRGs) with axons that project in the damaged nerve trunk. Macrophages and T-lymphocytes invade these ganglia where they are believed to release cytokines that lead to hyperexcitability and ectopic discharge, possibly contributing to neuropathic pain. Here, we examined the role of the sympathetic innervation in the inflammation of L5 DRGs of Wistar rats following transection of the sciatic nerve, comparing the effects of specific surgical interventions 10-14 days prior to the nerve lesion with those of chronic administration of adrenoceptor antagonists. Immunohistochemistry was used to define the invading immune cell populations 7 days after sciatic transection. Removal of sympathetic activity in the hind limb by transecting the preganglionic input to the relevant lumbar sympathetic ganglia (ipsi- or bilateral decentralization) or by ipsilateral removal of these ganglia with degeneration of postganglionic axons (denervation), caused less DRG inflammation than occurred after a sham sympathectomy. By contrast, denervation of the lymph node draining the lesion site potentiated T-cell influx. Systemic treatment with antagonists of α1-adrenoceptors (prazosin) or β-adrenoceptors (propranolol) led to opposite but unexpected effects on infiltration of DRGs after sciatic transection. Prazosin potentiated the influx of macrophages and CD4(+) T-lymphocytes whereas propranolol tended to reduce immune cell invasion. These data are hard to reconcile with many in vitro studies in which catecholamines acting mainly via β2-adrenoceptors have inhibited the activation and proliferation of immune cells following an inflammatory challenge.

Processing of central and reflex vagal drives by rat cardiac ganglion neurones: an intracellular analysis

Cardiac vagal tone is an important indicator of cardiovascular health, and its loss is an independent risk factor for arrhythmias and mortality. Several studies suggest that this loss of vagal tone can occur at the cardiac ganglion but the factors affecting ganglionic transmission in vivo are poorly understood. We have employed a novel approach allowing intracellular recordings from functionally connected cardiac vagal ganglion cells in the working heart-brainstem preparation. The atria were stabilised in situ preserving their central neural connections, and ganglion cells (n = 32) were impaled with sharp microelectrodes. Cardiac ganglion cells with vagal synaptic inputs (spontaneous, n = 10; or electrically evoked from the vagus, n = 3) were identified as principal neurones and showed tonic firing responses to current injected to their somata. Cells lacking vagal inputs (n = 19, presumed interneurones) were quiescent but showed phasic firing responses to depolarising current. In principal cells the ongoing action potentials and EPSPs exhibited respiratory modulation, with peak frequency in post-inspiration. Action potentials arose from unitary EPSPs and autocorrelation of those events showed that each ganglion cell received inputs from a single active preganglionic source. Peripheral chemoreceptor, arterial baroreceptor and diving response activation all evoked high frequency synaptic barrages in these cells, always from the same single preganglionic source. EPSP amplitudes showed frequency dependent depression, leading to more spike failures at shorter inter-event intervals. These findings indicate that rather than integrating convergent inputs, cardiac vagal postganglionic neurones gate preganglionic inputs, so regulating the proportion of central parasympathetic tone that is transmitted on to the heart.

On the Number of Preganglionic Neurons Driving Human Postganglionic Sympathetic Neurons: A Comparison of Modeling and Empirical Data

Postganglionic sympathetic axons in awake healthy human subjects, regardless of their identity as muscle vasoconstrictor, cutaneous vasoconstrictor, or sudomotor neurons, discharge with a low firing probability (∼30%), generate low firing rates (∼0.5 Hz) and typically fire only once per cardiac interval. The purpose of the present study was to use modeling of spike trains in an attempt to define the number of preganglionic neurons that drive an individual postganglionic neuron. Artificial spike trains were generated in 1–3 preganglionic neurons converging onto a single postganglionic neuron. Each preganglionic input fired with a mean interval distribution of either 1000, 1500, 2000, 2500, or 3000 ms and the SD varied between 0.5×, 1.0×, and 2.0× the mean interval; the discharge frequency of each preganglionic neuron exhibited positive skewness and kurtosis. Of the 45 patterns examined, the mean discharge properties of the postganglionic neuron could only be explained by it being driven by, on average, two preganglionic neurons firing with a mean interspike interval of 2500 ms and SD of 5000 ms. The mean firing rate resulting from this pattern was 0.22 Hz, comparable to that of spontaneously active muscle vasoconstrictor neurons in healthy subjects (0.40 Hz). Likewise, the distribution of the number of spikes per cardiac interval was similar between the modeled and actual data: 0 spikes (69.5 vs 66.6%), 1 spike (25.6 vs 21.2%), 2 spikes (4.3 vs 6.4%), 3 spikes (0.5 vs 1.7%), and 4 spikes (0.1 vs 0.7%). Although some features of the firing patterns could be explained by the postganglionic neuron being driven by a single preganglionic neuron, none of the emulated firing patterns generated by the firing of three preganglionic neurons matched the discharge of the real neurons. These modeling data indicate that, on average, human postganglionic sympathetic neurons are driven by two preganglionic inputs.

Ganglionic transmission in a vasomotor pathway studied in vivo

Intracellular recordings were made in vivo from 40 spontaneously active cells in the third lumbar sympathetic ganglion of urethane-anaesthetized rats. In 38/40 cells ongoing action potentials showed strong cardiac rhythmicity (93.4 +/- 1.9% modulation) indicating high barosensitivity and probable muscle vasoconstrictor (MVC) function. Subthreshold excitatory postsynaptic potentials (EPSPs) showed the same pattern. The 38 barosensitive neurons fired action potentials at 2.9 +/- 0.3 Hz. All action potentials were triggered by EPSPs, most of which were unitary events. Calculations indicated that <5% of action potentials were triggered by summation of otherwise subthreshold EPSPs. 'Dominant' synaptic inputs with a high safety factor were identified, confirming previous work. These were active in 24/38 cells and accounted for 32% of all action potentials; other ('secondary') inputs drove the remainder. Inputs (21 dominant, 19 secondary) attributed to single preganglionic neurons fired at 1.38 +/- 0.16 Hz. An average of two to three preganglionic neurons were estimated to drive each ganglion cell's action potentials. When cells were held hyperpolarized to block spiking, a range of spontaneous EPSP amplitudes was revealed. Threshold equivalent was defined as the membrane potential value that was exceeded by spontaneous EPSPs at the same frequency as the cell's original firing rate. In 10/12 cells examined, a continuum of EPSP amplitudes overlapped threshold equivalent. Small changes in cell excitability could therefore raise or lower the percentage of preganglionic inputs triggering action potentials. The results indicate that vasoconstrictor ganglion cells in vivo mostly behave not as 1:1 relays, but as continuously variable gates.

Synaptic transmission in the inferior mesenteric ganglion of hypertensive and normotensive rats

The inferior mesenteric ganglion (IMG) is a prevertebral sympathetic ganglion that innervates the lower abdominal and pelvic viscera. While extensive studies have been performed on the IMG of several species, there is very little information available concerning the rat IMG. Because of the importance of the mesenteric circulation in the regulation of systemic blood pressure and in the genesis of salt‐sensitive hypertension, we compared the properties and response profiles to pre‐ganglionic input of IMG neurons in normotensive (NT) and hypertensive (HT) animals using intracellular recording in vitro. All rats were uninephrectomized 4 weeks prior to experiments. HT rats received 200mg/kg of deoxycorticosterone s.q. and 1%NaCl/0.2%KCl in drinking water. NT animals (mean arterial pressure &lt; 130mmHg) received tap water. Resting membrane potential, rheobasic current, action potential amplitude, input resistance, and membrane time constant show no significant differences between NT and HT animals. In contrast to the findings in the IMG of other species, nearly all neurons (94% in HT, 91% in NT rats) showed a phasic response to direct current injection. This is the first study of the IMG of the hypertensive rat. While there is a predominance of phasic neurons in this ganglion, changes in sympathetic outflow that occur with hypertension are not due to alterations in the basic properties of the ganglionic neurons. Supported by NIH grant HL70687

Synaptic and membrane properties of parasympathetic ganglionic neurons innervating mouse trachea and bronchi

The pathophysiology of airway diseases, such as asthma, is increasingly studied using transgenic mice and other mouse models of airway inflammation where allergen-induced changes in airway smooth muscle tone and mucous secretion is due, in part, to activation of preganglionic airway parasympathetic nerves. Ganglionic parasympathetic neurons located in the airways in several species, including humans, have anatomical and electrophysiological properties that limit transmission of preganglionic synaptic input. In this study, intracellular recordings were made from neurons in parasympathetic ganglia located on the trachea and bronchi of adult mice to determine electrophysiological properties associated with regulation of transmission of preganglionic input. Ganglionic neurons were characterized as having either tonic or phasic action potential accommodation patterns. Tonic neurons responded with repetitive action potentials sustained throughout a depolarizing current step, whereas phasic neurons generated one or a burst of action potential(s) and accommodated. A small subset displayed both patterns. Phasic neurons could be further differentiated as usually having either short- or long-duration afterhyperpolarizing potential following single and multiple action potentials. In most cells, stimulation of preganglionic nerves elicited one population of nicotinic fast excitatory postsynaptic potentials that were graded in amplitude, usually suprathreshold for action potential generation, and did not decrease in amplitude during higher frequency stimulation. Dye injection into the neurons revealed that dendrites were either absent or very short. These results provide evidence that in contrast to the characteristics of airway parasympathetic neurons reported in other species, including human, the electrophysiological and synaptic properties, and anatomical characteristics of mouse lower airway ganglionic neurons, are less associated with integration of presynaptic input.

Anatomical and physiological properties of pelvic ganglion neurons in female mice

Most neurons that regulate motility and blood flow in female pelvic organs are located within pelvic (paracervical) ganglia. In this study we investigated the anatomical and physiological properties of neurons within mouse (C57/Bl/6) paracervical ganglia. Most neurons showed immunoreactivity for choline acetyl transferase (CHAT) and were presumably cholinergic. Few neurons (~ 5%) were tyrosine hydroxylase (TH) positive. Immunohistochemical labelling for microtubule associated protein 2 showed most neurons had small somata (cross sectional area ~ 300 µm 2) and lacked dendrites. Action potential (AP) discharge characteristics, determined by depolarising current step injection, revealed most neurons (70%) adapted rapidly to depolarising current injection and were classified as “phasic”. The remaining neurons discharged APs throughout the current step and were classified as “tonic”. Membrane properties and current–voltage relationships were similar in phasic and tonic neurons, however the afterhyperpolarisation was significantly smaller in tonic neurons. Stimulation of preganglionic axons usually evoked a single strong preganglionic input (21/27 and 9/10 for pelvic and hypogastric nerves, respectively). In 19 preparations where we tested for inputs from both nerves pelvic inputs predominated (23/45 neurons) and inputs via the hypogastric nerve were rarely observed (3/45 neurons). Together, our data indicate that most neurons within mouse paracervical ganglia are cholinergic and parasympathetic. As there is little anatomical or functional evidence for integration of preganglionic inputs we propose that the role of paracervical neurons is restricted to one of spatial amplification or filtering of preganglionic inputs.

Cell-Autonomous Inhibition of α7-Containing Nicotinic Acetylcholine Receptors Prevents Death of Parasympathetic Neurons during Development

Neurotrophic molecules are key retrograde influences of cell survival in the developing nervous system, but other influences such as activity are also emerging as important factors. In the avian ciliary ganglion, half the neurons are eliminated between embryonic day 8 (E8) and E14, but it is not known how cell death is initiated. Because systemic application of alpha7-nicotinic acetylcholine receptor (nAChR) antagonists prevents this cell loss, we examined differences in receptor densities and responses of intracellular calcium to nicotine using the calcium-sensitive dye fura-2. In addition, we determined whether cell-autonomous inhibition of alpha7 activation in neurons prevented cell death. E8 neurons are heterogeneous with respect to alpha7-nAChR density, which leads to large increases in [Ca2+]i in some neurons; E8 neurons also exhibit a slower rate of Ca2+ decay after nicotinic stimulation than E13 neurons. Expressing alpha-bungarotoxin that is tethered to the membrane by a glycosylphosphatidylinositol linkage (GPIalpha btx) in ciliary ganglion neurons with the retroviral vector RCASBP(A) blocks increases in intracellular calcium induced by nicotine through alpha7-nAChRs and prevents neurons from dying. Expression of GPIalpha btx in surrounding non-neural tissues, but not in neurons, does not prevent cell loss. Furthermore, the GPIalpha btx is not efficiently expressed in the accessory oculomotor neurons, eliminating preganglionic inputs as another site for action of the antagonist. These results support the hypothesis that cholinergic inputs facilitate cell death in the developing autonomic nervous system by activating alpha7-nAChRs, possibly by leading to increases in intracellular calcium that exceed the threshold for cell survival.

Synaptic transmission in sympathetic vasoconstrictor pathways and its modification after injuries

The discharge of vasoconstrictor pathways arising in the CNS is largely unmodified as it passes through the sympathetic ganglia to the vasculature. The underlying synaptic events have been revealed by intracellular recordings from sympathetic paravertebral ganglion cells in the course of ongoing and reflex activity in anesthetized animals, first made in Skok’s Laboratory in Kyiv (Ukraine). Each preganglionic neuron diverges to contact a number of post-ganglionic neurons, on each of which several pre-ganglionic inputs converge. However, only suprathreshold “strong,” or “dominant” synapses are effective in transmitting the CNS signals. Strong synapses differ from the other subthreshold “weak,” or “accessory” inputs: (a) excitatory synaptic currents are >1 nA in their amplitude, (b) 3 to ≈>30 times more quanta of acetylcholine are released, (c) pre-synaptic Ca2+ entry through channels resistant to all-known antagonists triggers acetylcholine release, and (d) post-synaptic Ca2+ entry boosts and prolongs the nicotinic current. While the majority of postganglionic neurons have only one strong input, a proportion receives two or, rarely, three such inputs. In cells with multiple strong inputs, an equivalent number of discrete Ca2+ currents can be evoked at distinct foci electrically distant from the soma, suggesting that each strong input has a unique dendritic association with a cluster of Ca2+ channels. When strong preganglionic inputs are destroyed, residual weak synapses sprout and rapidly restore the suprathreshold connections. While much remains to be discovered about how strong synapses are established, their high safety factor ensures the wide and secure distribution of vasoconstrictor command signals from the CNS.

Diversity of sympathetic vasoconstrictor pathways and their plasticity after spinal cord injury

Sympathetic vasoconstrictor pathways pass through paravertebral ganglia carrying ongoing and reflex activity arising within the central nervous system to their vascular targets. The pattern of reflex activity is selective for particular vascular beds and appropriate for the physiological outcome (vasoconstriction or vasodilation). The preganglionic signals are distributed to most postganglionic neurones in ganglia via synapses that are always suprathreshold for action potential initiation (like skeletal neuromuscular junctions).Most postganglionic neurones receive only one of these “strong” inputs, other preganglionic connections being ineffective. Pre- and postganglionic neurones discharge normally at frequencies of 0.5–1 Hz and maximally in short bursts at <10 Hz. Animal experiments have revealed unexpected changes in these pathways following spinal cord injury. (1) After destruction of preganglionic neurones or axons, surviving terminals in ganglia sprout and rapidly re-establish strong connections, probably even to inappropriate postganglionic neurones. This could explain aberrant reflexes after spinal cord injury. (2) Cutaneous (tail) and splanchnic (mesenteric) arteries taken from below a spinal transection show dramatically enhanced responses in vitro to norepinephrine released from perivascular nerves. However the mechanisms that are modified differ between the two vessels, being mostly postjunctional in the tail artery and mostly prejunctional in the mesenteric artery. The changes are mimicked when postganglionic neurones are silenced by removal of their preganglionic input. Whether or not other arteries are also hyperresponsive to reflex activation, these observations suggest that the greatest contribution to raised peripheral resistance in autonomic dysreflexia follows the modifications of neurovascular transmission.

Structure of peripheral synapses: autonomic ganglia

Final motor neurons in sympathetic and parasympathetic ganglia receive synaptic inputs from preganglionic neurons. Quantitative ultrastructural analyses have shown that the spatial distribution of these synapses is mostly sparse and random. Typically, only about 1%-2% of the neuronal surface is covered with synapses, with the rest of the neuronal surface being closely enclosed by Schwann cell processes. The number of synaptic inputs is correlated with the dendritic complexity of the target neuron, and the total number of synaptic contacts is related to the surface area of the post-synaptic neuron. Overall, most neurons receive fewer than 150 synaptic contacts, with individual preganglionic inputs providing between 10 and 50 synaptic contacts. This variation is probably one determinant of synaptic strength in autonomic ganglia. Many neurons in prevertebral sympathetic ganglia receive additional convergent synaptic inputs from intestinofugal neurons located in the enteric plexuses. The neurons support these additional inputs via larger dendritic arborisations together with a higher overall synaptic density. There is considerable neurochemical heterogeneity in presynaptic boutons. Some synapses apparently lack most of the proteins normally required for fast transmitter release and probably do not take part in conventional ganglionic transmission. Furthermore, most preganglionic boutons in the ganglionic neuropil do not form direct synaptic contacts with any neurons. Nevertheless, these boutons may well contribute to slow transmission processes that need not require conventional synaptic structures.