Changes in the synaptic activity in sciatic nerve-extensor digitorum longus (preparations induced by ethanol)

The aim of this research is the study of the modification of synaptic activity caused by ethanol in the rat sciatic nerve-extensor digitorum longus (EDL) muscle preparation. For such a purpose, intracellular recordings have been carried out, keeping the muscle immersed in normal Ringer solution and in Ringer solutions containing ethanol at different concentrations up to 0,8 M. Therefore, the resting potential of muscle cells and the frequency of m.e.p.p.s were measured. Qualitative observations of m.e.p.p.s shape were also carried out. Ethanol increases the frequency of m.e.p.p.s in the rat sciatic nerve - EDL muscle preparation. The logarithm of relative frequency (frequency in Ringer solution with ethanol/frequency in normal Ringer solution) is linear with respect to the concentration of ethanol, with a slope of 1.44. Furthermore, ethanol increases the amplitude and lengthens the time course of m.e.p.p.s. The muscle cells undergo a hyperpolarization of about 2-3% at the lowest concentrations of ethanol tested.     

The recovery of motor innervation during treatment with ethanol

We have studied the effects of the fluidizing action of ethanol on motor reinnervation in order to clarify if membrane fluidity changes affect synaptic plasticity and nerve regeneration. Sprague Dawley rats were denervated by crushing the sciatic nerve and subsequently, on the 16th day, the degree of reinnervation of the EDL muscle was observed by electrophysiological technique: in particular an observation was made of the resting potential and the m.e.p.p.s frequency by intracellular recordings in muscle fibers. During the nerve regeneration period, the rats were treated with 3 g of ethanol per kilogram of body weight per day. We have found that ethanol quickens the resting potential recovery but does not affect the m.e.p.p.s frequency.     

4-aminoquinoline-induced 'giant' miniature endplate potentials at mammalian neuromuscular junctions

4-Aminoquinoline (4-AQ) in concentrations around 200 micrometers induces, within minutes of its application to isolated mouse or rat neuromuscular junctions, the appearance of a population of miniature endplate potentials (m.e.p.ps) with a larger than normal amplitude, so-called giant m.e.p.ps (g.m.e.p.ps). With amplitudes 2-12 times the modal value of m.e.p.p. amplitude, the population of g.m.e.p.ps varied between 15 and 45% of the total population of m.e.p.ps. There was no increase in the frequency of m.e.p.ps but a positive correlation between the frequency of g.m.e.p.ps and the total frequency of m.e.p.ps. In many instances the rise time and decay time of g.m.e.p.ps were prolonged compared to normal. Elevated extracellular calcium concentrations increased the frequency of m.e.p.ps but had no effect on g.m.e.p.p. frequency. High extracellular potassium concentrations markedly increased m.e.p.p. frequency but failed to influence g.m.e.p.p. frequency. Similar observations were made with ethanol 0.1 M, ouabain 200 micrometers or black widow spider venom. Botulinum toxin type A markedly reduced total m.e.p.p. frequency but 4-AQ still induced g.m.e.p.ps. Nerve stimulation failed to release quanta corresponding to the g.m.e.p.ps. G.m.e.p.ps seemed to originate from quantal acetylcholine release from the nerve terminal since they were abolished by surgical denervation and by the addition of d-tubocurarine to the medium. Blockade of voltage-sensitive calcium or sodium channels by, respectively, manganese ions or tetrodotoxin failed to affect the appearance and the frequency of g.m.e.p.ps. The electrophysiological findings and a statistical analysis of the characteristics of the m.e.p.ps indicate that they belong to two populations. One population is accelerated by the depolarization-release coupling mechanism responsible for evoked transmitter release and is characterized by an amplitude distribution and a process in time that indicate that they correspond to releases occurring at 'active zones' in the nerve terminal. The second population of m.e.p.ps is uninfluenced by nerve terminal depolarization and transmembrane calcium fluxes. This population apparently originates from sites dispersed in the nerve terminal membrane and outside the 'active zones'. 4-AQ increases the frequency of this second m.e.p.p. population without affecting the first population.     

Evidence for nicotinic acetylcholine receptors on nasal trigeminal nerve endings of the rat

The peripheral chemoreceptors of the trigeminal system in the nasal cavity are presumed to be free nerve endings arising from Adelta and C fibers. These fibers appear to be scattered throughout the nasal epithelium, and arise from the nasopalatine and ethmoid branches of the trigeminal nerve. In the present study, the effects of nicotinic acetylcholine receptor (nAChR) blockers on ethmoid nerve responses to nicotine and cyclohexanone were examined. Multiunit neural recordings were obtained from the ethmoid nerve of Sprague-Dawley rats. Vapor-phase nicotine (12.5 p.p.m.) and cyclohexanone (450 p.p. m.) were delivered to the rats' nares via an air-dilution olfactometer. The magnitude of the response to nicotine decreased after the administration of the nAChR blockers dihydro-beta-erythroidine hydrobromide (DHBE) and mecamylamine hydrochloride. DHBE is a competitive nicotinic receptor antagonist specific for the alpha4beta2 receptor subtype and mecamylamine is known to bind alpha3beta4 and alpha4beta2 receptors. The nAChR blockers had no effect on ethmoid nerve responses to cyclohexanone. These results suggest that the mechanism by which at least one irritant stimulates nasal trigeminal nerve endings involves the binding of irritant with a specific receptor.     

Capsaicin Modifies Responses of Rat Chorda Tympani Nerve Fibers to NaCl

Single-fiber preparations of the rat chorda tympani (CT) nervewere used to study the mechanism of action of capsaicin on salt-tastetransduction. Capsaicin selectively suppressed the responsesto NaCl of the CT nerve fibers (N-fibers) that are sodium-specific(insensitive or poorly sensitive to potassium). Among the morebroadly responsive, cation-sensitive fibers (E-fibers) thereare two subtypes, both of which responded to capsaicin but indifferent ways (‘enhanced’ type and ‘suppressed’type). In both N- and E-fibers, 5% ethanol (the vehicle forcapsaicin) slightly reduced the response to 100 mM NaCl. Thesuppressive effect of capsaicin on the response of the N-typefibers to 100 mM NaCl was significantly stronger than the effectof 5% ethanol. The suppression lasted for at least 20 s afterthe simultaneous application of 100 p.p.m. capsaicin-100 mMNaCl. These results indicate that 100 p.p.m. capsaicin can modifythe response of CT fibers to NaCl. The observed effect of capsaicinon gustatory fibers could be the net result of opposite suppressiveand enhancing processes in the taste buds cells and excitedintra- or extragemmal trigeminal nerve endings. Chem. Senses22: 249–255, 1997. ^*These authors contributed equally to this study     

Post-tetanic potentiation of miniature end-plate potential frequency at neuromuscular junction of the rat soleus muscle

1. Changes in miniature end-plate potential (m.e.p.p.) frequency by repetitive nerve stimulation were examined in the rat soleus muscle. 2. The increase of m.e.p.p. frequency was induced by repetitive stimulation and persisted for several minutes after the tetanus. That is, post-tetanic potentiation (PTP) of neuromuscular transmission was first demonstrated here in the rat soleus muscle. 3. The time course of the decay of m.e.p.p. frequency after the tetanus showed a double exponential curve which consisted of a fast decaying component (augmentation) and a slow decaying component (potentiation). 4. The magnitude of PTP depended on the stimulation frequency and its duration. It increased with the increase of duration and was at its maximum at a frequency of 100 Hz. 5. No PTP was elicited by repetitive stimulation under conditions in which end-plate potential (e.p.p.) was completely suppressed, and, moreover, m.e.p.p. frequency tended to decrease after the tetanus.     

Action of botulinum A toxin and tetanus toxin on synaptic transmission

Intracellular recordings of the spontaneous activity from mammalian spinal cord neurons in culture demonstrated different sensitivities of excitatory and inhibitory synaptic transmission for the action of tetanus toxin (Tetx) and botulinum toxin type A (Botx). The effects of Tetx and Botx on spontaneous and nerve-evoked transmitter release were compared under identical experimental conditions in experiments on in vitro poisoned mouse diaphragms. At 37 degrees C completely paralyzed endplates are characterized by a very low frequency of spontaneous miniature endplate potentials (m.e.p.p.s) and by a 100% failure to evoke endplate potentials (e.p.p.s) in response to single nerve stimuli. Striking differences in the action of both toxins have been observed when the very low transmitter release probabilities of paralyzed nerve-muscle preparations were increased by tetanic nerve stimulation and/or application of potent K+-channel blockers and/or by reduction of temperature to 25 degrees C. While Botx did not change the short latency between nerve impulse and postsynaptic response, Tetx produced a temporal dispersion of the quantal release suggesting that the toxins act at different sites in the chain of events that result in transmitter release. To find further evidence to support the different actions of the toxins the spontaneous transmitter release was studied in more detail. Tetx blocked preferentially the release of so-called large mode m.e.p.p.s without affecting the frequency of the small mode ones. In contrast, Botx strongly inhibited both the small and large mode m.e.p.p.s.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of prostaglandins E1 and F2 alpha on neuromuscular transmission in the frog sartorius muscle.

End plate potentials (e.p.p.s.) and miniature end plate potentials (m.e.p.p.s.) were recorded intracellularly at the neuromuscular junction of the frog sartorius muscle. Addition of as little as 8.5 x10(-8)M PGE1 reduced the mean m.e.p.p. frequency. The mean amplitude of m.e.p.p.s was not changed, the mean amplitude of the e.p.p.s and the quantum content of the transmitter released by a nerve impulse was slightly reduced. A decrease in mean m.e.p.p. frequency was also seen in response to the administration of 8.5 x 10(-8)M PG2 alpha. The mean amplitude of e.p.p.s and m.e.p.p.s and the quantum content remained unchanged. The possible presynaptic mode of action of PGs in the preparation of discussed.     

Transmitter release in botulinum-poisoned muscles

Examination of miniature end-plate potentials (m.e.p.ps) in rat skeletal muscle poisoned in vivo by botulinum toxin type A reveals the presence of two populations of potentials. One population which corresponds to m.e.p.ps in unpoisoned muscles and to quantal end-plate potentials. The frequency of these m.e.p.ps is greatly reduced by botulinum toxin. The second population of m.e.p.ps has quite different characteristics. These m.e.p.ps have a more variable, but generally much larger amplitude, and their time to peak is longer than normal m.e.p.ps. The frequency of these m.e.p.ps increases during poisoning and reaches 0.3-1 Hz after 10-14 days. In addition to the variability in amplitude and time-to-peak these m.e.p.ps differ from those at unpoisoned junctions by being unaffected by procedures which alter extra- or intracellular Ca2+ concentrations. The appearance of this Ca2+-insensitive spontaneous quantal secretion of acetylcholine is apparently not a direct effect of the toxin but secondary to blockade of impulse transmission since it also appears at unpoisoned end-plates when transmission is impaired for other reasons. Procedures which increase the intracellular Ca2+ concentration in nerve terminals restore transmitter release from botulinum toxin poisoned nerves. Furthermore, the block caused by the toxin is very temperature-dependent, a reduction in temperature relieving the block. Since presynaptic Ca2+ currents are unaltered by the toxin it is proposed that the block of transmission is due to a reduction in the calcium content of the nerve terminal to a level where the amount of Ca2+, which normally enters, is insufficient to activate transmitter release.     

Cyclic nucleotides and neuromuscular transmission.

A review of the research on cyclic nucleotides and neuromuscular transmission suggests that cAMP is involved in the release of transmitter from motor nerve endings. Lipid-soluble derivations of cAMP cause depolarization of unstimulated nerve endings and prolong the after potentials of stimulated nerve endings. They also increase the frequency of miniature end plate potentials and increase the quantal content of stimulus evoked end plate potentials. Similar effects are produced by compounds that activate adenylate cyclase or inhibit phosphodiesterase. The responses to the derivatives of cAMP and activators of cyclase are enhanced by inhibitors of phosphodiesterase and prevented by compounds that block the flux of calcium into nerve endings. There is no evidence that suggests that cyclic nucleotides are involved in the postjunctional response to transmitter. Thus, it seems likely that cAMP is involved in the regulation of calcium in motor nerve endings and the exocytosis of transmitter. Additional study should expand our knowledge of neuromuscular transmission and contribute to an understanding of the functions of cyclic nucleotides in other synapses.     

MICROTUBULE PROTEIN : Identification in and Transport to Nerve Endings

The subunit protein of microtubules, tubulin, has been demonstrated to be present in isolated nerve endings by gel electrophoresis, amino acid composition, and peptide mapping. The tubulin constitutes approximately 28% of the soluble protein of the nerve endings. The transport of tubulin to the nerve endings has been demonstrated and its relationship to slow transport is discussed.     

Supravital Methylene Blue Staining of Piloneural Complexes of Common Fur Hair Follicles in the Rat

Light microscopic observations employing supravital methylene blue staining are presented for piloneural complexes of common fur hairs in the mystacial pad of the rat snout. The investigation revealed anatomical details of piloneural complexes belonging to follicles of both vellus and guard hairs. In the methylene blue stained preparations, different types of palisade-like lanceolate nerve fiber endings could be discriminated. The thicker vellus and thinner guard hairs (hair diameter: 15-25 μm) exhibited a different innervation pattern compared to the thicker guard hairs, and two subtypes of piloneural complexes could be distinguished. Both subtypes were characterized by slightly stained lanceolate endings and the absence of a circular nerve fiber plexus. One subtype, however, showed strongly stained spines originating from the lanceolate endings. A few spines of adjacent lanceolate endings appeared in contact with each other. In the second subtype, these spines were replaced by anastomoses suggesting a delicate terminal nerve fiber network. The moderately stained lanceolate endings located primarily at the follicles of thicker guard hairs (hair diameter: 30-40 μm) showed smooth outlines, but were characterized by the occurrence of an intensely stained additional circular nerve fiber plexus. The differences in the morphology of piloneural complexes associated with the follicles of common fur hairs suggest differences regarding their mechanoreceptive tasks.     

Spontaneous activity of the isolated rabbit superior cervical sympathetic ganglion

Investigation of spontaneous activity (mean amplitude of spikes 200–300 µV, frequency from 0.07 to 2.9 Hz) in the rabbit superior cervical sympathetic ganglion by the sucrose gap method showed that this activity was completely blocked by D-tubocurarine and hexamethonium; its frequency was increased in hypertonic solution, by an increase in the external potassium concentration, and by the addition of theophylline and ethanol. These observations suggest that the activity observed is due to spontaneous liberation of acetylcholine mediator from preganglionic nerve endings. However, addition of tetrodotoxin and an increase in the external calcium concentration to 10 mM block spontaneous activity in the ganglion. This suggests that the observed spontaneous activity consists of action potentials.     

Supravital Methylene Blue Staining of Piloneural Complexes of Common Fur Hair Follicles in the Rat

Light microscopic observations employing supravital methylene blue staining are presented for piloneural complexes of common fur hairs in the mystacial pad of the rat snout. The investigation revealed anatomical details of piloneural complexes belonging to follicles of both vellus and guard hairs. In the methylene blue stained preparations, different types of palisade-like lanceolate nerve fiber endings could be discriminated. The thicker vellus and thinner guard hairs (hair diameter: 15-25 μm) exhibited a different innervation pattern compared to the thicker guard hairs, and two subtypes of piloneural complexes could be distinguished. Both subtypes were characterized by slightly stained lanceolate endings and the absence of a circular nerve fiber plexus. One subtype, however, showed strongly stained spines originating from the lanceolate endings. A few spines of adjacent lanceolate endings appeared in contact with each other. In the second subtype, these spines were replaced by anastomoses suggesting a delicate terminal nerve fiber network. The moderately stained lanceolate endings located primarily at the follicles of thicker guard hairs (hair diameter: 30-40 μm) showed smooth outlines, but were characterized by the occurrence of an intensely stained additional circular nerve fiber plexus. The differences in the morphology of piloneural complexes associated with the follicles of common fur hairs suggest differences regarding their mechanoreceptive tasks.     

Histochemical study on the nerve endings in the lung of Rattus rattus rufescens and Francolinus pondicerianus

Presence and the relation of the nerve endings with associated structures in the lund of Rattus rattus rufescens (Indian black rat) and Francolinus pondicerianus (grey partridge or safed teeter) has been studied by cholinesterase technique. Dot, plate, and Vater pacini Corpuscles like nerve endings in the lung of Rattus and dot, and bulb like nerve endings with axis cylinder covered with myelinated sheath in the lung of Francolinus were recorded. The nerve endings were AChE-positive.     

The influence of hindlimb unloading on the effectiveness of modulation of the mediator secretion via the autoreceptor system

In experiments on neuromuscular synapses of rat fast (m. Extensor digitorum longus, EDL) and slow (m. soleus) skeletal muscles, changes in the intensity of spontaneous quantal mediator secretion in response to the activation of presynaptic cholinoreceptors by the nonhydrolyzable acetylcholine analogue carbachol and to an increase in K+ concentration in the control group of animals and in animals subjected to different terms of unloading of hindlimbs have been compared. The intensity of spontaneous secretion of mediator quanta was evaluated from the mean frequency of miniature endplate potentials. In the control group of animals, the frequency of miniature endplate potentials by the action of carbachol increased by 363% in m. EDL and by 62% in m. soleus. The frequency of miniature endplate potentials in the synapses of m. EDL was more sensitive to K(+)-induced depolarization too. The bearing unloading of hindlimbs abolished the sensitivity of spontaneous secretion to carbachol in the synapses of m. EDL, whereas in m. soleus it was unchanged. However, the preservation of sensitivity of nerve endings of fast muscle to K(+)-induced depolarization allows one to assume that the hindlimb unloading leads to a decrease in the number of functioning presynaptic receptors.     

Quantitative evaluation of sensory nerve endings in hypertrophy of labia minora pudendi in women.

The authors studied the types of sensory nerve endings in hypertrophic labia minora pudendi in women aged between 18 and 69. The nervous tissue was impregnated with silver nitrate by Lavrentyev's modification of the Bielschowski-Gross method. The sensory nerve endings were divided into the following groups: free endings and arborizations, spray-like endings, seven types of clew-like nerve endings, and Pacinian corpuscles. It is the clew-like endings that absolutely predominate, they were 2,027 in number. The arborizations and spray-like endings found were 105 in number and only 4 of Pacinian corpuscles were found. As compared with normal labia there are no principal differences among the types of sensory nerve endings. The main difference consists in the rate of occurrence (89.6 as against 57.8) and in the thickness of the capsule, while the nerve clew proper does not grow in diameter. Therefore, we are of theopinion that a more frequent occurrence and larger size of the capsule is associated with local changes in the organ (especially with hypertrophy of the connective tissue). We observed no differences due to age either in the occurrence of the individual types or in the thickness of the capsule. Great individual variability in both directions was seen. The findings corroborated again the rare occurrence of meissner's endings in the labia minora in woman. Therefore it may be assumed that it is rather a random finding and that it is a type of clew-like nerve ending. Finally, we presume that the individually varying rate of occurrence of the nerve endings in the external genitals could also affect general sensitivity in this region, including sexual sensitivity.     

Discrepancies between spontaneous and evoked synaptic potentials at normal, regenerating and botulinum toxin poisoned mammalian neuromuscular junctions

Amplitudes and times to peak of spontaneous miniature endplate potentials (m.e.p.ps) and evoked quantal endplate potentials (e.p.ps) were compared at normal, regenerating and botulinum toxin poisoned neuromuscular junctions of the extensor digitorum longus muscle of the rat. At normal junctions the mean time to peak of m.e.p.ps was longer and more variable than that of similar-sized e.p.ps. At endplates where nerve regeneration was induced by mechanical crushing of the motor nerve the frequency of m.e.p.ps was reduced and their amplitude distribution was broader than normal. The distribution of times to peak of m.e.p.ps was considerably broader than that of quantal e.p.ps recorded at the same endplates. At neuromuscular junctions poisoned with botulinum toxin type A, spontaneous and evoked transmitter release were greatly reduced. The amplitude distribution of m.e.p.ps was wider than that of e.p.ps and the time to peak of e.p.ps was about twice as fast as and less variable than that of m.e.p.ps. To explain the observed differences in time to peak among m.e.p.ps and between m.e.p.ps and quantal e.p.ps we suggest that some m.e.p.ps, but not e.p.ps, originate from transmitter quanta released from sites at a greater distance from postsynaptic receptors or that the release or diffusion process for acetylcholine is more prolonged when producing some of the m.e.p.ps. Such mechanisms produce at normal junctions a small population of m.e.p.ps with prolonged times to peak, at regenerating junctions a greater proportion of such m.e.p.ps and in botulinum toxin poisoning a majority.     

Identification of Different Types of Spinal Afferent Nerve Endings That Encode Noxious and Innocuous Stimuli in the Large Intestine Using a Novel Anterograde Tracing Technique

In mammals, sensory stimuli in visceral organs, including those that underlie pain perception, are detected by spinal afferent neurons, whose cell bodies lie in dorsal root ganglia (DRG). One of the major challenges in visceral organs has been how to identify the different types of nerve endings of spinal afferents that transduce sensory stimuli into action potentials. The reason why spinal afferent nerve endings have been so challenging to identify is because no techniques have been available, until now, that can selectively label only spinal afferents, in high resolution. We have utilized an anterograde tracing technique, recently developed in our laboratory, which facilitates selective labeling of only spinal afferent axons and their nerve endings in visceral organs. Mice were anesthetized, lumbosacral DRGs surgically exposed, then injected with dextran-amine. Seven days post-surgery, the large intestine was removed. The characteristics of thirteen types of spinal afferent nerve endings were identified in detail. The greatest proportion of nerve endings was in submucosa (32%), circular muscle (25%) and myenteric ganglia (22%). Two morphologically distinct classes innervated myenteric ganglia. These were most commonly a novel class of intraganglionic varicose endings (IGVEs) and occasionally rectal intraganglionic laminar endings (rIGLEs). Three distinct classes of varicose nerve endings were found to innervate the submucosa and circular muscle, while one class innervated internodal strands, blood vessels, crypts of lieberkuhn, the mucosa and the longitudinal muscle. Distinct populations of sensory endings were CGRP-positive. We present the first complete characterization of the different types of spinal afferent nerve endings in a mammalian visceral organ. The findings reveal an unexpectedly complex array of different types of primary afferent endings that innervate specific layers of the large intestine. Some of the novel classes of nerve endings identified must underlie the transduction of noxious and/or innocuous stimuli from the large intestine.     

Innervation of mouse lymph nodes: nerve endings on muscular vessels and reticular cells

Lymph node nerve endings have been studied in 1- to 48-day-old mice. Serial sections of Epon-embedded lymph nodes were observed under the electron microscope to find the nerve endings. Most lymph node nerve fibers finally reach the smooth muscle cells of arterioles and muscular venules. Both kinds of vascular endings are similar, although endings are less numerous on venules. Nerve endings consist of one or more nerve processes surrounded by a usually incomplete Schwann cell sheath; frequently, axons show wide areas directly facing the muscle cells. The distance between such a naked axon and a myocyte ranges from 100 to 800 nm. Small granulated and clear vesicles are especially abundant in varicosities of nerve processes that are located very close to muscle cells. Nerve endings of lymph node vasculature probably correspond to vasomotor sympathetic adrenergic endings, regulating the degree of contraction of vessels which have a muscular layer. Other kinds of nerve endings also exist in lymph nodes: some axons appear free in the stroma and contact the surfaces of reticular cells; the latter also extend delicate cytoplasmic processes that surround the axons. The functional significance of nerve cell-reticular cell contacts is unknown.