Preganglionic Splanchnic Nerve Research Articles

Cholinergic and peptidergic neurotransmission in the adrenal medulla: A dynamic control of stimulus-secretion coupling.

Synaptic neurotransmission at the splanchnic nerve-chromaffin cell synapse is a chief element of the stimulus-secretion coupling in the adrenal medullary tissue, managing and regulating the secretion of catecholamines. Making the state of play more intricate than initially envisioned, the synaptic vesicles of nerve terminals innervating the medulla contain various compounds, including various neurotransmitters and neuropeptides. Under basal conditions associated with a low splanchnic nerve discharge rate, neurotransmission is ensured by the synaptic release of the primary neurotransmitter acetylcholine (ACh). Under sustained and repetitive stimulations of the splanchnic nerve, as triggered in response to stressors, the synaptic release of neuropeptides, such as the pituitary adenylate cyclase-activating polypeptide PACAP, supplants ACh release. The anatomical and functional changes that occur presynaptically at the preganglionic splanchnic nerve, combined with changes occurring postsynaptically at nicotinic acetylcholine receptors (nAChRs), confer the adrenomedullary synapses a solid and persistent aptitude to functional remodeling, from birth to aging. The present review focuses on the composite cholinergic and noncholinergic nature of neurotransmission occurring at the splanchnic nerve-chromaffin cell synapse and its remodeling in response to physiological or pathological stimuli.

Sigma-1 receptor ligands inhibit catecholamine secretion from adrenal chromaffin cells due to block of nicotinic acetylcholine receptors.

Adrenal chromaffin cells (ACCs) are the neuroendocrine arm of the sympathetic nervous system and key mediators of the physiological stress response. Acetylcholine (ACh) released from preganglionic splanchnic nerves activates nicotinic acetylcholine receptors (nAChRs) on chromaffin cells causing membrane depolarization, opening voltage-gated Ca2+ channels (VGCC), and exocytosis of catecholamines and neuropeptides. The serotonin transporter is expressed in ACCs and interacts with 5-HT1A receptors to control secretion. In addition to blocking the serotonin transporter, some selective serotonin reuptake inhibitors (SSRIs) are also agonists at sigma-1 receptors which function as intracellular chaperone proteins and can translocate to the plasma membrane to modulate ion channels. Therefore, we investigated whether SSRIs and other sigma-1 receptor ligands can modulate stimulus-secretion coupling in ACCs. Escitalopram and fluvoxamine (100nM to 1μM) reversibly inhibited nAChR currents. The sigma-1 receptor antagonists NE-100 and BD-1047 also blocked nAChR currents (≈50% block at 100nM) as did PRE-084, a sigma-1 receptor agonist. Block of nAChR currents by fluvoxamine and NE-100 was not additive suggesting a common site of action. VGCC currents were unaffected by the drugs. Neither the increase in cytosolic [Ca2+ ] nor the resulting catecholamine secretion evoked by direct membrane depolarization to bypass nAChRs was altered by fluvoxamine or NE-100. However, both Ca2+ entry and catecholamine secretion evoked by the cholinergic agonist carbachol were significantly reduced by fluvoxamine or NE-100. Together, our data suggest that sigma-1 receptors do not acutely regulate catecholamine secretion. Rather, SSRIs and other sigma-1 receptor ligands inhibit secretion evoked by cholinergic stimulation because of direct block of Ca2+ entry via nAChRs.

The cardiac-related rhythm in preganglionic sympathetic activities of developing piglets

This investigation was performed to determine whether partial spectral analysis of preganglionic sympathetic nerve discharges would reveal age-related differences in the distribution of baroreceptor afferent information to brainstem sympathetic-related neurons. Any influence of baroreceptor afferent activity on ordinary spectra of cervical sympathetic and splanchnic nerves was removed by partialization using the arterial blood pressure signal which represented baroreceptor activity. An absence of statistically significant coherence in partialized nerve spectra would indicate that sympathetic-related neurons receive peripheral baroreceptor afferent input, but are not interconnected, whereas the presence of significant coherence would mean that these neurons are interconnected. Ordinary spectral analysis did not demonstrate age-related differences in the relationship between nerve activity and baroreceptor afferent input. In many animals, large peaks, located at cardiac frequencies (range 2.75–5.6 Hz), were noted in ordinary nerve autopower spectra, and were significantly correlated in ordinary coherence spectra. Partialization of nerve spectra eliminated or reduced cardiac-related peaks in autopower spectra regardless of age, and, in 8 of 10 animals, reduced coherence estimates to non-significant values. In two animals, 19 and 36 days old, significant coherence values remained after partialization. These results demonstrated that cardiac-related peaks in coherence in spectra of preganglionic splanchnic and cervical sympathetic nerves were dependent upon peripheral afferent baroreceptor input in most animals. Further, the finding that significant residual coherence was absent in most cases suggested a paucity of intrabulbar pathways connecting brainstem sympathetic-related neurons.

Effects of Ca2+ channel antagonists on acetylcholine and catecholamine releases in the in vivo rat adrenal medulla

To elucidate the types of voltage-dependent Ca(2+) channels controlling ACh and catecholamine releases in the in vivo adrenal medulla, we implanted microdialysis probes in the left adrenal medulla of anesthetized rats and investigated the effects of Ca(2+) channel antagonists on ACh, norepinephrine, and epinephrine releases induced by nerve stimulation. The dialysis probes were perfused with Ringer solution containing a cholinesterase inhibitor, neostigmine. The left splanchnic nerves were electrically stimulated at 2 and 4 Hz before and after intravenous administration of Ca(2+) channel antagonists. omega-Conotoxin GVIA (an N-type Ca(2+) channel antagonist, 10 microg/kg) inhibited ACh release at 2 and 4 Hz by approximately 40%, norepinephrine release at 4 Hz by approximately 50%, and epinephrine release at 2 and 4 Hz by approximately 45%. A fivefold higher dose of omega-conotoxin GVIA (50 microg/kg) did not further inhibit these releases. omega-Conotoxin MVIIC (a P/Q-type Ca(2+) channel antagonist, 50 microg/kg) inhibited ACh and epinephrine releases at 4 Hz by approximately 30%. Combined omega-conotoxin GVIA (50 microg/kg) and MVIIC (250 microg/kg) inhibited ACh release at 2 and 4 Hz by approximately 70% and norepinephrine and epinephrine releases at 2 and 4 Hz by approximately 80%. Nifedipine (an L-type Ca(2+) channel antagonist, 300 and 900 microg/kg) did not change ACh release at 2 and 4 Hz; however, nifedipine (300 microg/kg) inhibited epinephrine release at 4 Hz by 20%, and nifedipine (900 microg/kg) inhibited norepinephrine and epinephrine releases at 4 Hz by 30%. In conclusion, both N- and P/Q-type Ca(2+) channels control ACh release on preganglionic splanchnic nerve endings while L-type Ca(2+) channels do not. L-type Ca(2+) channels are involved in norepinephrine and epinephrine releases on chromaffin cells.

Exotoxin-insensitive G proteins mediate synaptically evoked muscarinic sodium current in rabbit sympathetic neurones.

1. The involvement of G proteins in the transduction pathway that links muscarinic receptors to the low-threshold voltage-dependent sodium current (INa,M) was studied in neurones from intact sympathetic prevertebral ganglia using the whole-cell configuration of the patch-clamp technique. Experiments were performed in the presence of the nicotinic receptor antagonists hexamethonium (50 microM) and d-tubocurarine (50 microM). 2. INa,M was activated by either bath-applying muscarinic agonists or stimulating the preganglionic splanchnic nerves. Synaptically and agonist-mediated INa,M did not display significant run-down or changes in their properties in cells tested, irrespective of whether the pipette solutions contained GTP. 3. Dialysis of sympathetic neurones with GDP beta S (500-750 microM) decreased the amplitude of INa,M by approximately 65% compared with control neurones within 30 min. 4. In the absence of muscarinic receptor stimulation, intracellular dialysis with GTP gamma S (500 microM) for 10 min slowly and slightly (20-25%) activated INa,M. GTP gamma S dialysis markedly slowed down the decay of INa,M after its transient activation with carbachol pulses (10-20 s) or nerve stimulation (3-5 s). The INa,M activation became fully irreversible 2.9 min after the start of GTP gamma S dialysis. Dialysing cells with the G protein activator AIF4-led to a rapid but transient activation of INa,M. 5. Synaptically and agonist-evoked INa,M were not affected in ganglia treated with 0.5-1 microgram ml-1 pertussis toxin (PTX) for 7-24 h at 37 degrees C. Control experiments showed that this treatment severely reduced the PTX-sensitive inhibition of N-type calcium currents induced by carbachol (CCh) and noradrenaline. Application of NEM (N-ethylmaleimide) for 2 min depressed the INa,M evoked in response to bath-applied CCh by only 27%. 6. Incubating ganglia with 5-10 micrograms ml-1 of cholera toxin for 7 h had no effect on the carbachol-induced INa,M but greatly potentiated (approximately 250%) the synaptically evoked INa,M, presumably via a presynaptic mechanism. 7. These results show that the coupling between muscarinic receptors and NaM channels is mediated by pertussis toxin- and cholera toxin-insensitive G proteins, possibly of the Gq/11 or G12 class.

Differentiated sympathetic neural control of the kidney.

Anatomic and neurophysiological methods were used to identify functionally specific subgroups of renal sympathetic nerve fibers. The distribution of diameters of the predominating unmyelinated fibers showed a major mode at 1.1 microns and a minor mode at 1.6 microns. The conduction velocity was 2.10 +/- 0.10 m/s, consistent with unmyelinated C fibers. Analysis of strength-duration relationships during renal nerve stimulation showed that both rheobase and chronaxie values for renal blood flow were greater than those for urinary flow rate and were independent of stimulation frequency. This difference suggests a higher stimulation threshold (smaller diameter) for those renal nerve fibers involved in the renal blood flow response (renal vasoconstriction) compared with those for the urinary flow rate response (antidiuresis) to renal nerve stimulation. Single renal units that responded to preganglionic splanchnic nerve stimulation were studied. Those with spontaneous activity (88%) responded to stimulation of arterial baroreceptors, arterial and central chemoreceptors, and peripheral thermoreceptors, whereas those that lacked spontaneous activity (12%) responded only to stimulation of peripheral thermoreceptors (known to produce renal vasoconstriction). A minority population of single renal units has been identified that, although renal vasoconstrictor, does not exhibit other characteristic features of vasoconstrictor neurons (i.e., responsiveness to stimulation of arterial baroreceptors and arterial and central chemoreceptors). These findings suggest the existence of functionally specific subgroups of renal nerve fibers.

Cytophotometric and electrophysiological analysis of the role of the celiac plexus in maintenance of temperature homeostasis during cold stress in rats

The role of the celiac plexus in maintaining temperature homeostasis in rats during cold stress was studied histochemically, ultrastructurally, and electrophysiologically. Inhibition on efferent impulses in preganglionic (splanchnic) nerves and facilitation in postganglionic (superior mesenteric) nerves during a short-term cold exposure leading to slight hypothermia were found. In cold-exposed animals after the celiac plexus decentralization, the neurons showed an increase in the fluorescence intensity and activity of energy metabolism enzymes, hyperplasia of fine structures responsible for protein synthesis and energy supply of cells. It is suggested that during cold stress when efferent impulse flow in preganglionic nerves markedly decreases, the celiac plexus becomes a regulatory centre for autonomic functions involved in the maintenance of temperature homeostasis.

Inhibition of sympathetic nerve activity by acute administration of the tricyclic antidepressant desipramine

In rats anesthetized with pentobarbital/chloral hydrate, the i.v. injection of desipramine (0.3 mg/kg) caused a rapid fall in mean arterial blood pressure accompanied by a reduction in efferent renal nerve integrated activity. A similar reduction in electrical activity was seen in the preganglionic splanchnic nerve. Pretreatment of the rats with yohimbine (0.3 mg/kg) increased renal nerve activity by 20%, and the decrease in blood pressure and renal nerve activity elicited by a subsequent dose of desipramine was blocked. I.v. injection of prazosin (0.05 mg/kg) caused a reduction in systemic blood pressure. The hypotensive effect of a subsequent dose of desipramine was largely blocked, whereas the fall in renal nerve activity was attenuated, but still present. The data are consistent with an action of desipramine to reduce central sympathetic tone by increasing the release of endogenous noradrenaline which acts on α 2-adrenoceptors at CNS sites controlling sympathetic activity. An additional involvement of central α 1-adrenoceptors cannot be ruled out.

Selective control of sympathetic pathways to the kidney, spleen and intestine by the ventrolateral medulla in rats.

1. Electrical activity of multifibre renal, splenic, mesenteric and greater splanchnic nerves and 13th thoracic white rami was recorded in artificially respired, urethane-anaesthetized rats. Discharge of neurones in the rostral ventrolateral medulla was blocked by unilateral microinjections of the inhibitory amino acid glycine and effects on the electrical activity of these sympathetic nerves were compared. 2. Blockade of the rostral ventrolateral medulla caused greater decreases in discharge of renal than splenic nerves and had no consistent effect on mesenteric nerves. This blockade also decreased the discharge of the preganglionic white rami more than that of the preganglionic splanchnic nerves. 3. Postganglionic responses to rostral ventrolateral medulla blockade were always greater than preganglionic responses. 4. The arterial pressure and renal nerve responses to rostral ventrolateral medulla blockade in urethane-anaesthetized rats were not different from those in rats anaesthetized with alpha-chloralose. 5. These findings demonstrate that pre- and postganglionic sympathetic pathways to the kidney are more dependent upon excitatory drive from the rostral ventrolateral medulla than pathways directed to the spleen and intestine.

OPIOID RECEPTOR MODULATION OF NEURAL TRANSMISSION IN THE RABBIT COELIAC GANGLION AND GANGLIONIC OPIOID RECEPTOR ACTIVATION BY BUNITROLOL

1. We examined the preganglionic splanchnic nerve activity and postganglionic renal nerve activity before and after a local injection of naloxone (20 micrograms/kg) into the coeliac ganglion of anaesthetized rabbits. This was done during graded hypertension, induced by the administration of phenylephrine (0.5-10 micrograms/kg, i.v.) and with selective intraganglionic injection of methionine-enkephalin (ME) and bunitrolol, which is a beta-blocker. 2. During hypertension both pre- and postganglionic discharge decreased, but only postganglionic discharge was inhibited by naloxone treatment into the ganglion. 3. Local injection of ME (0.1-10 micrograms/kg) into the coeliac ganglion decreased postganglionic activity by 9.0 +/- 1.0 to 41.2 +/- 4.7% from control, and this decrease was inhibited by naloxone. 4. Administration of bunitrolol (1-300 micrograms/kg) decreased postganglionic discharge by 3.9 +/- 1.4 to 39.7 +/- 2.4% of the control and this decrease was also inhibited by naloxone. 5. These results suggest that opioid receptors in the coeliac ganglion play an inhibitory role in neural ganglionic transmission and that this inhibitory action reduces postganglionic sympathetic discharge.

Activation of mesenteric arteries and veins by preganglionic and postganglionic nerves.

Intracellular recordings were made from inferior mesenteric artery and vein and their primary and secondary branches. The major nerve trunks associated with the inferior mesenteric ganglion were stimulated with single shocks and repetitively. Excitatory junction potentials (EJPs) were recorded in arterial cells but not in veins. Stimulation of the postganglionic lumbar colonic nerves elicited the largest EJPs; stimulation of preganglionic lumbar splanchnic nerves or the interganglionic intermesenteric nerves elicited EJPs that were 40 and 44%, respectively, of the amplitude of the EJPs elicited with lumbar colonic nerve stimulation. In response to repetitive stimulation, the membranes of both artery and vein cells depolarized by an amount dependent on the frequency and duration of stimulation. At a given frequency of stimulation, the vein cells depolarized more rapidly and constricted sooner than the arterial cells. The depolarizations and the associated constrictions were attenuated by phentolamine (10(-6) M) or prazosin (10(-6) M). In 70% of arterial cells tested, repetitive nerve stimulation at 1-5 Hz resulted in a long-lasting hyperpolarization. At 2 Hz the hyperpolarizations averaged 2.0 +/- 0.4 mV. These hyperpolarizations were not blocked by either alpha- or beta-adrenergic antagonists. These studies demonstrate that the innervation of mesenteric arteries and veins can be activated by stimulating preganglionic, interganglionic, and postganglionic nerves and that there are fundamental differences between the electrical responses of adjacent arteries and veins to nerve stimulation.

Animal pharmacology of guanfacine

The pharmacologic data obtained from animal experiments with guanfacine, a novel, centrally acting antihypertensive agent, are reviewed. When given orally, guanfacine lowers systemic blood pressure in conscious DOCA-NaCl-hypertensive rats, Grollman rats and spontaneously hypertensive rats in a dose-dependent manner. It is also effective in renal hypertensive cats. Guanfacine reduces blood pressure in cats, rabbits and rats after injection into the lateral cerebral ventricle and in dogs after infusion into the vertebral artery at intravenously ineffective doses. Vagally mediated reflex bradycardia in dogs is enhanced. The preganglionic splanchnic (sympathetic) nerve activity is reduced in cats. In rats, guanfacine reduces the noradrenaline turnover in the brain stem. All these findings indicate a central site of action. Peripheral α-adrenoceptor stimulant properties of guanfacine have been demonstrated in various studies. In addition to postsynaptic stimulant effects, presynaptic guanfacine-induced inhibition of sympathetic heart nerve stimulation is antagonized by rauwolscine but not by prazosin, indicating a highly preferential α 2-agonistic presynpatic action of the drug. In receptor binding studies using rat cortex membranes and human platelets, guanfacine exhibited a high selectivity for α 2 adrenoceptors. Guanfacine has the advantage over other centrally acting antihypertensives of being less sedative and causing no rebound hypertension after discontinuation of treatment. The latter is mainly due to its pharmacokinetic properties.

Role of sympathetic nerve activity in endotoxin induced hypotension in cats.

Within 10 min of intravenous (iv) injection of E. coli endotoxin (1 mg X kg-1) in alpha-chloralose anaesthetised cats, mean blood pressure (MBP) fell significantly to 86% of the pre-endotoxin level. There followed a gradual decline in MBP, so that after 60 min MBP was depressed to 49% of the pre-endotoxin level. Preganglionic splanchnic nerve (PSN) activity decreased significantly before the rapid fall in MBP. There followed a gradual decrease in PSN activity coincident with the significant fall in MBP, so that 60 min after iv injection of E. coli endotoxin PSN activity fell to 40% of the pre-endotoxin level. Heart rate did not differ significantly from the pre-endotoxin level in the control group. However, the hypotension and the reduction of PSN activity following endotoxin were abolished by intracisternal (ic) pretreatment with phentolamine (0.5 mg X kg-1). The heart rate in this group increased significantly, by 15% at 10 min and was maintained until the end of the experiment. These results indicate that in E. coli endotoxin hypotension the blood pressure falls together with the reduction in sympathetic outflow; and suggests that stimulation of central alpha-adrenergic receptors leads to an inhibition of activity in brainstem sympathetic pathways.

Neural control of sympathetic nerve response to cardiogenic hypotension in anesthetized cat.

The present study was designed to determine effect of the preganglionic splanchnic nerve activity (SNA) on the brief hypotension accompanied with the occlusion of left circumflex coronary artery (CxCAO) in chloralose anesthetized cats. Following CxCAO in animals with neuraxis intact, no significant alterations of SNA occurred despite the significant fall in mean blood pressure (MBP). A significant fall in MBP also occurred in vagotomized animals with arterial baroreceptors intact, but SNA was significantly augmented from 12.9 +/- 2.7 impulses/sec before CxCAO to 24.4 +/- 4.3 impulses/sec 60 sec after the occlusion. In vagotomized animals, in which their carotid sinuses were isolated and perfused with the constant pressure at a level equal to systemic blood pressure (112 +/- 6 mmHg) and with higher pressure (167 +/- 7 mmHg), SNA was not altered significantly during the hypotension due to CxCAO. When the carotid sinuses were perfused with lower pressure (53 +/- 8 mmHg), a significant increase in SNA occurred simultaneously with the decrease in MBP after CxCAO. The peak decreases in blood pressure during the coronary occlusion were significantly greater in the vagotomized group (-46 +/- 5 mmHg) and in the Low-CSP group (-50 +/- 5 mmHg) than in other groups. Onset of this excitatory efferent sympathetic response to the hypotension due to the coronary occlusion in the vagotomized and Low-CSP groups was delayed significantly despite a significant fall in arterial blood pressure. These results show that vagal afferents from the heart may play a role of inhibiting the sympathetic augmentation mediated by arterial baroreceptors during cardiogenic hypotension. An excessive activation of cardiac receptors with sympathetic afferents may be induced by the profound fall in blood pressure, resulting in further impairment of cardiac function due to progressive myocardial ischemia under the condition of high sympathetic tone activated by baroreceptor reflex.

Effect of naloxone on baroreflex, sympathetic tone and blood pressure in the cat

Studies were designed to determine the effect of naloxone on sympathetic nerve activity and blood pressure and, also to investigate the central effect of naloxone in relation to the baroreflex system in α-chloralose-anesthetized cats. Following intravenous injection of naloxone (2 mg/kg), preganglionic splanchnic nerve (PSN) activity significantly increased in parallel to the increase in blood pressure. Set gain (mmHg −1) of carotid sinus baroreflex together with the operational range (mmHg) was increased after naloxone. Intraventricular (4th) injection of naloxone (0.2 mg/kg) produced identical responses in blood pressure and PSN activity as well as an altered set gain and range of the baroreflex. Additionally, the pressor response to carotid hypotension and medullary pressor area stimulation were augmented during naloxone activation but not the pressor response to posterior hypothalamus stimulation. The depressor responses to stimulation of both carotid sinus nerve and medullary depressor region were facilitated by naloxone. These data suggest that naloxone has an effect on the central autonomic blood pressure regulatory circuits which participate in the carotid sinus baroreflex system.

Centrally mediated cardiovascular effects of B-HT 920 (6-allyl-2-amino-5,6,7,8-tetrahydro-4H-thiazolo-[4,5-d]-azepine dihydrochloride), a hypotensive agent of the "clonidine type".

Intravenous injection of 30 micrograms/kg of B-HT 920 (6-allyl-2-amino-5,6,7,8-tetrahydro-4H-thiazolo-[4,5-d]-azepine dihydrochloride) into cats lead initially to an increase in blood pressure and then to a long-lasting decrease in blood pressure and heart rate. The central site of the hypotensive and bradycardiac action was demonstrated by the significantly greater effect after intracisternal (i.ci.) than after intravenous injection of B-HT 920, 3 micrograms/kg. The drug decreased the rate of spontaneous discharges in preganglionic splanchnic nerve fibers of normal and noradrenaline-depleted cats (3 micrograms/kg, i.ci.). Vagally mediated reflex bradycardia elicited by angiotensin injection in beta-adrenoceptor-blocked dogs was facilitated by intracisternal injection of 10 micrograms/kg B-HT 920. Both sympathoinhibition and vagal reflex facilitation were antagonized by the alpha-adrenoceptor-blocking agent piperoxan (50 micrograms/kg, i.ci). Therefore, B-HT 920 can be classified as an agent of the clonidine type, despite its different chemical structure. Quantitative differences between B-HT 920 and clonidine are discussed with respect to the greater alpha 2/alpha 1 adrenoceptor activity ratio of the former drug, as reported previously.

Effect of timolol on autonomic neural discharge associated with ouabain-induced arrhythmia

The purpose of this study was to determine the effect of the β-blocking agent timolol on postganglionic cardiac and preganglionic splanchnic sympathetic and vagal neural discharge, ouabain-induced arrhythmia, heart rate and mean arterial blood pressure. Cats were anesthetized with α-chloralose, given atropine, and pretreated with timolol (5 mg/kg, i.v. infused at a rate of 0.5 mg/kg/min for 10 min). Bolus injections of ouabain (25 μg/kg, i.v.) were given every 15 min until the animals died; the first injection was given 15 min after the end of the timolol infusion. When compared with cats (n = 13) receiving only ouabain, timolol (n = 11 cats) increased the time to ouabain-induced arrhythmia and death from 23 ± 3 to 48 ± 7 and 46 ± 3 to 76 ± 9 min, respectively (P < 0.05). Both heart rate and mean arterial blood pressure fell immediately after the onset of the timolol infusion. Eleven minutes later, the mean heart rate and blood pressure had decreased from 137 ± 3 to 104 ± 6 beats/min and 133 ± 6 to 103 ± 7 mm Hg, respectively (P < 0.05); ouabain did not reverse the decreases. Neural activity monitored from the vagus and from the postganglionic cardiac and preganglionic splanchnic sympathetic nerves was not significantly altered by the infusion of timolol. The administration of ouabain after timolol did not alter splanchnic nor vagal discharge. Most important was the observation that postganglionic cardiac sympathetic neural discharge exhibited both increases and decreases, i.e., a nonuniform neural discharge, at the time of ouabain-induced arrhythmia and that the ouabain-induced nonuniformity did not occur in the cats pretreated with timolol. Thus, the protective effect of timolol may be due, in part, to prevention of the nonuniform postganglionic cardiac sympathetic neural discharge and to the prevention of ouabain-induced increases in vagal discharge. The establishment of β-blockade and a direct negative inotropic action may also have contributed to the antiarrhythmic action of timolol.

Competitive interaction between foreign nerves innervating frog skeletal muscle.

1. Competition between two foreign nerves innervating frog skeletal muscle has been studied by using pairs of somatic motor nerves (s.m.n.s) or one s.m.n. and the preganglionic splanchnic nerve (s.p.n.) implanted into a denervated sartorius muscle that has been transplanted to the lymph sac of the back. 2. A single s.m.n. implanted into the muscle succeeded in innervating essentially every fibre within 2--3 months; tetanic stimulation of the nerve elicited 9--100% of the maximal direct tetanus tension. Most of the e.p.p.s were suprathreshold, since a single indirect stimulus evoked a twitch 60--100% as large as that to a direct stimulus. 3. If two s.m.n.s were implanted simultaneously, tetanic stimulation of either elicited 80--100% of the maximal tension to direct stimulation. If one nerve was implanted 2--3 months before the other, the second, although usually less effective than the first, normally innervated 50--100% of the fibres, with approximately the same time course of innervation as a single s.m.n. 4. Mutual synaptic repression was seen on examination of twitch tensions. With either simultaneous or staggered innervation, stimulation of each s.m.n. resulted in a twitch of 30--50% of the total direct twitch tension, with little overlap between the fields driven by the two nerves. Intracellular recordings showed that the distribution of subthreshold and spike-producing e.p.p.s reflected the existence of separate twitch fields. Even if one s.m.n. was implanted several months before the other and had time to establish suprathreshold junctions on most muscle fibres, an s.m.n. implanted later was able to reduce sharply the effectiveness of many junctions from the earlier nerve while itself innervating most muscle fibres. 5. The subthreshold e.p.p.s had low quantal content, typically ten or fewer quanta/e.p.p. The min e.p.p. frequency was very low, while min e.p.p. amplitude appeared to be normal. 6. In the vast majority of muscle fibres, junctions from the two nerves were not within recording distance of each other. Hence, we infer that the competitive interaction was mediated somehow via the muscle fibre. 7. The preganglionic splanchnic nerve, which also successfully reinnervated frog skeletal muscle, competed with a foreign s.m.n. in ways which differ qualitatively from the competition by a second s.m.n. In the presence of a s.m.n., synapses of the s.p.n. were almost universally subthreshold. However, if the s.p.n. was implanted 2--3 months before the s.m.n., the s.m.n. was prevented for several months from innervating fibres driven by the s.p.n. This delay in s.m.n. reinnervation was greater than if the first nerve implanted was also an s.m.n. 8. After 6--8 months of dual innervation by s.m.n. and s.p.n., the s.m.n. became almost totally dominant. However, if the s.m.n. was then sectioned, the s.p.n. became as effective, within approximately 1 week, as it would have been in the absence of the s.m.n.

The physiology, pharmacology, and trophic effectiveness of synapses formed by autonomic preganglionic nerves on frog skeletal muscle.

1. Frog sartorius muscles, newly denervated and transplanted to the lymph sac of the back, are reinnervated by implanted cholinergic nerves (spinal somatic motor nerves or the preganglionic sympathetic splanchnic nerve), but not by nerves). 2. Foreign somatic motor nerves (s.m.n.s) form synapses that resemble normal sartorius neuromuscular junctions electrophysiologically. 3. Axons of the sympathetic preganglionic splanchnic nerve (s.p.n.) grow throughout the muscle, but only a small percentage of fibres form synapses. Most e.p.p.s are of low quantal content, generally subthreshold. Long onset latencies and multiple post-synaptic responses indicate that innervation is multiple, multi-terminal, and by unmyelinated axons. 4. Spontaneous miniature e.p.p.s at splanchnic junctions occur at an average rate under 0.1/sec. Their average amplitude and time course are about the same as for control muscles, but the variability of amplitudes is greater than for control muscles. 5. The amount of facilitation shown by s.p.n.-evoked e.p.p.s is the same as by s.m.n. e.p.p.s, but the time course is almost twice as long. 6. S.p.n.-reinnervated fibres show dramatic post-tetanic potentiation preceded by depression, following as few as 20--50 stimuli. 7. As judged by standard physiological and histochemical criteria, AChEsterase is absent at s.p.n. junctions. 8. The pharmacological responses of the s.p.n. junctions are similar to those of normal or foreign s.m.n. innervated neuromuscular junctions in their sensitivity to the cholinergic blocking agents D-tubocurarine and hexamethonium. 9 The s.p.n. is capable of restricting ACh sensitivity to the sites of nerve contacts, although this restriction occurs more slowly and less completely than with s.m.n. reinnervation. The loss of extrajunctional ACh sensitivity can be correlated with effectiveness of innervation; but significant restriction occurs even in s.p.n. reinnervated fibres that probably never contract to nerve stimulation.

Pharmacological characterization of B-HT 933 (2-amino-6-ethyl-4,5,7,8,-tetrahydro-6H-oxazolo-[5,4-d]-azepindihydrochloride) as a hypotensive agent of the "clonidine-type".

The hypotensive action of a new compound, B-HT 933 (2-amino-6-ethyl-4,5,7,8-tetrahydro-6H-oxazolo-[5,4-d]-azepin) was characterized to be of the “clonidine-type” by the use of 6 pharmacological test procedures. (1) Intravenous injection of the substance elicited a typical cardiovascular reaction pattern of initial hypertension, followed by hypotension with bradycardia and decreased cardiac output. (2) A direct α-adrenoceptor stimulation at peripheral vascular sites was demonstrated by a hypertensive action in spinal rats. (3) A central nervous site of action was shown as 30 μg/kg injected intracisternally (i.ci.) lowered blood pressure and heart rate in cats, but the same dose i.v. was practically ineffective. (4) The decrease in sympathetic activity was revealed by a decrease in preganglionic splanchnic nerve discharges in cats. (5) The drug facilitated the vagally-mediated baroreceptor reflex as shown by the increase in reflex bradycardia following i.v. angiotensin injection in dogs. (6) The involvement of central nervous α-adrenoceptors was suggested because the α-adrenoceptor blocking drug piperoxan antagonized the facilitation of the baroreceptor reflex induced by B-HT 933. Furthermore, the drug facilitated the flexor reflex in rats with spinal section at midthoracic level. These results were unexpected as the chemical structure of B-HT 933 differs considerably from hitherto known substances of the “clonidine-type”.