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S C Landis

Publications and source records attributed to S C Landis.

At least 73 records · Page 4Linked to original sources
PubMed1990

Asynchronous appearance and topographic segregation of neuropeptide-containing cells in the developing rat adrenal medulla.

The developmental expression of neuropeptide Y (NPY) and leucine-enkephalin (L-Enk) was examined in embryonic, early postnatal, and adult chromaffin cells with double- and triple-label immunocytochemical techniques and compared to the expression of immunoreactivity for tyrosine hydroxylase (TH) and phenylethanolamine N-methyltransferase (PNMT). In addition, the establishment of preganglionic innervation was assessed by labeling for choline acetyltransferase (ChAT) and L-Enk. NPY-IR was detectable on embryonic (E) day 15 in a clustered subpopulation of TH-IR cells. L-Enk and PNMT-IR cells were initially present on E16 in a separate nonclustered population of TH-IR cells. By late embryonic development, twice as many TH-IR cells expressed NPY and 4 times as many expressed L-Enk as in the adult. In contrast to early embryonic development, NPY-IR was evident in both the clustered and nonclustered subpopulation of TH-IR cells at this time. The proportion of NPY-IR chromaffin cells decreased to adult values during the first postnatal week at the time when obviously clustered TH-IR cells were no longer observed. The embryonic rise in the proportion of L-Enk-IR cells correlates with the developmental increase in glucocorticoid production, while the postnatal decrease corresponds to the appearance of ChAT-IR in the preganglionic innervation of the adrenal medulla. These results indicate that NPY and L-Enk are expressed at different times and in different subpopulations of cells in the embryonic adrenal. Further, the observation that peptide expression by chromaffin cells undergoes marked changes during development raises the possibility that a number of factors including developmental history, environmental signals and impulse activity play a role in the regulation of neuropeptide expression in sympathoadrenal derivatives of the neural crest.

Adrenal Medulla↗
PubMed1989

Substance P and neurokinin A in the cat carotid body: localization, exogenous effects and changes in content in response to arterial pO2.

In the present work we studied the occurrence of substance P (SP) and neurokinin A (NKA) in the carotid bodies of cats by means of immunocytochemistry and radioimmunoassay (RIA). We also compared the exogenous effects of SP and NKA on carotid body sensory discharge. SP- and NKA-like immunoreactivities (SP-LI; NKA-LI) were seen in many glomus cells and in a sparse plexus of fine fibers. The SP-LI containing glomus cells and fibers also exhibited NKA-LI, suggesting that both these tachykinins coexist in the carotid body. Chemoreceptor discharge increased both by SP and NKA in a dose-dependent manner. The peak excitation produced by SP and NKA was the same when the effects were compared on an equimolar basis. The tachykinin content of the carotid bodies varied with changes in arterial pO2. During normoxia, SP and NKA levels were 57 +/- 8 and 85 +/- 14 fmol/mg, respectively. When the animals were exposed for 1 h to 100% O2, SP content was unchanged (51 +/- 4 fmol/mg), whereas NKA levels were significantly lower than during normoxia (29 +/- 3 fmol/mg, P less than 0.01). Following 1 h of hypoxia, SP content of the carotid body was 146 +/- 20 fmol/mg, a value higher than that obtained during normoxia and hyperoxia. NKA levels, on the other hand, were not significantly different from normoxic values. These results indicate that the cat carotid body (1) contains both SP and NKA, (2) both peptides augment neural discharge of the carotid body and (3) their levels in the carotid body are substantially altered by arterial oxygen, the natural stimulus to the chemoreceptors.

Action Potentials↗
PubMed1989

Differentiation of noradrenergic traits in the principal neurons and small intensely fluorescent cells of the parasympathetic sphenopalatine ganglion of the rat.

Catecholamine synthetic enzymes are found in many cranial parasympathetic principal neurons, and in the small intensely fluorescent (SIF) cells that populate parasympathetic as well as sympathetic ganglia. While there is evidence that the acquisition of noradrenergic properties in sympathetic neuron precursors depends on factors that these cells encounter in the trunk environment, the mechanisms that direct the development of noradrenergic traits in cranial parasympathetic neurons and SIF cells are not understood. The present study examines the time course of appearance of tyrosine hydroxylase (TH) immunoreactivity in the principal neurons and SIF cells of the rat sphenopalatine ganglion. We show that the sphenopalatine ganglion of normal adult rats contains both a small population of TH-immunoreactive principal neurons and many SIF cells. The TH-immunoreactive principal neurons do not synthesize or store detectable catecholamines, even though the majority of sphenopalatine ganglion neurons do contain 1-amino acid decarboxylase catalytic activity. Sphenopalatine ganglion principal neurons do not accumulate detectable levels of exogenous catecholamines. This observation suggests that they lack a high affinity norepinephrine uptake system. In contrast to what has been observed previously for sympathetic neurons, the appearance of TH immunoreactivity in sphenopalatine neurons is not temporally correlated with the cessation of neural crest cell migration. The first TH-immunoreactive neurons do not appear in the sphenopalatine ganglion until Embryonic Day 16.5, 2 days after the ganglion has condensed and process outgrowth has begun. The number of sphenopalatine neurons that express TH immunoreactivity increases dramatically between Embryonic Day 18.5 and Postnatal Day 1, but then decreases. In fact, the percentage of sphenopalatine neurons that express TH immunoreactivity is almost fivefold higher in newborn than in adult rats. SIF cells cannot be definitively identified in the sphenopalatine ganglion until after Embryonic Day 18.5. The time course of appearance of TH immunoreactivity in sphenopalatine ganglion cells raises the possibility that TH expression is stimulated in these cells by factors encountered either at their condensation site or at their target, such as glucocorticoids or nerve growth factor. The relatively late appearance of SIF cells in the sphenopalatine ganglion argues against the hypothesis that SIF cells are the precursors of all autonomic neurons.

Animals↗
PubMed1988

Cholinergic phenotype developed by noradrenergic sympathetic neurons after innervation of a novel cholinergic target in vivo.

Mammalian sympathetic neurons in vivo may express either a noradrenergic or cholinergic phenotype. In view of the opposing effect of noradrenaline and acetylcholine on most autonomic target organs, the target-appropriate expression of neurotransmitter is critical. We have examined the maturation of the sympathetic innervation of rat sweat glands to define the developmental mechanisms regulating neurotransmitter choice in vivo. Eccrine sweat glands and their sympathetic innervation develop together postnatally in the rat. Early postnatal innervation expresses only noradrenergic properties, but as the glands and their innervation mature, noradrenergic properties decrease dramatically and cholinergic features appear in the same population of neurons. To investigate the role of the sweat gland in this change we have used a transplantation paradigm which allows sweat glands to be innervated by sympathetic neurons that would normally innervate noradrenergic target organs and remain noradrenergic throughout life. We observe that the sympathetic neurons that innervate the novel cholinergic target alter their neurotransmitter properties and develop a cholinergic phenotype. These results indicate that target organs are able to induce appropriate neurotransmitter traits in the neurons that innervate them.

Acetylcholinesterase↗
PubMed1988

Evidence for neurotransmitter plasticity in vivo. II. Immunocytochemical studies of rat sweat gland innervation during development.

Previous studies of the cholinergic sympathetic innervation of rat sweat glands provide evidence for a change in neurotransmitter phenotype from noradrenergic to cholinergic during development. To define further the developmental history of cholinergic sympathetic neurons, we have used immunocytochemical techniques to examine developing and mature sweat gland innervation for the presence of the catecholamine synthetic enzymes tyrosine hydroxylase (TH) and dopamine beta-hydroxylase (DBH) and for two neuropeptides present in the mature cholinergic innervation, vasoactive intestinal peptide (VIP) and calcitonin gene-related peptide (CGRP). In 7-day old animals, intensely TH- and DBH-immunoreactive axons were closely associated with the forming glands. The intensity of both the TH and DBH immunofluorescence decreased as the glands and their innervation developed. Neither TH-IR nor DBH-IR disappeared entirely; faint immunoreactivity for both enzymes was reproducibly detected in mature animals. In contrast to noradrenergic properties, the expression of peptide immunoreactivities appeared relatively late. No VIP-IR or CGRP-IR was detectable in the sweat gland innervation at 4 or 7 days. In some glands VIP-IR first appeared in axons at 10 days, and was evident in all glands by 14 days. CGRP-IR was detectable only after 14 days. In addition to VIP-IR and CGRP-IR, we examined the sweat gland innervation for several neuropeptides which have been described in noradrenergic sympathetic neurons including neuropeptide Y, somatostatin, substance P, and leu- and met-enkephalin; these peptides were not evident in either developing or mature sweat gland axons. Our observations provide further evidence for the early expression and subsequent modulation of noradrenergic properties in a population of cholinergic sympathetic neurons in vivo. In addition, the asynchronous appearance during development of the two neuropeptide immunoreactivities raises the possibility that the expression of peptide phenotypes may be controlled independently.

Animals↗
PubMed1988

Developmental interactions between sweat glands and the sympathetic neurons which innervate them: effects of delayed innervation on neurotransmitter plasticity and gland maturation.

The neurotransmitter properties of the sympathetic innervation of sweat glands in rat footpads have previously been shown to undergo a striking change during development. When axons first reach the developing glands, they contain catecholamine histofluorescence and immunoreactivity for catecholamine synthetic enzymes. As the glands and their innervation mature, catecholamines disappear and cholinergic and peptidergic properties appear. Final maturation of the sweat glands, assayed by secretory competence, is correlated temporally with the development of cholinergic function in the innervation. To determine if the neurotransmitter phenotype of sympathetic neurons developing in vivo is plastic, if sympathetic targets can play a role in determining neurotransmitter properties of the neurons which innervate them, and if gland maturation is dependent upon its innervation, the normal developmental interaction between sweat glands and their innervation was disrupted. This was accomplished by a single injection of 6-hydroxy-dopamine (6-OHDA) on Postnatal Day 2. Following this treatment, the arrival of noradrenergic sympathetic axons at the developing glands was delayed 7 to 10 days. Like the gland innervation of normal rats, the axons which innervated the sweat glands of 6-OHDA-treated animals acquired cholinergic function and their expression of endogenous catecholamines declined. The change in neurotransmitter properties, however, occurred later in development than in untreated animals and was not always complete. Even in adult animals, some fibers continued to express endogenous catecholamines and many nerve terminals contained a small proportion of small granular vesicles after permanganate fixation. The gland innervation in the 6-OHDA-treated animals also differed from that of normal rats in that immunoreactivity for VIP was not expressed in the majority of glands. It seems likely that following treatment with 6-OHDA sweat glands were innervated both by neurons that would normally have done so and by neurons that would normally have innervated other, noradrenergic targets in the footpads, such as blood vessels. Contact with sweat glands, therefore, appears to suppress noradrenergic function and induce cholinergic function not only in the neurons which normally innervate the glands but also in neurons which ordinarily innervate other targets. Effects of delayed innervation were also observed on target development. The appearance of sensitivity to cholinergic agonists by the sweat glands was coupled with the onset of cholinergic transmission.(ABSTRACT TRUNCATED AT 400 WORDS)

Age Factors↗
PubMed1988

The ultrastructural distribution of putative nicotinic receptors on cultured neurons from the rat superior cervical ganglion.

The distribution of putative nicotinic receptors on cultured neurons from the rat superior cervical ganglion was determined by electron microscopic autoradiography using a radioactively labeled snake venom neurotoxin, toxin F. In a previous study, we demonstrated that toxin F blocks nicotinic transmission in these cultures of sympathetic neurons and in intact superior cervical ganglia. [125I]toxin F bound to two sites in these cultures: one site that was also recognized by the neuromuscular blocker, alpha-bungarotoxin, and a second site that was not. Since alpha-bungarotoxin neither blocks nicotinic transmission nor prevents the blocking effects of toxin F, the site specific to the binding of toxin F most probably represents neuronal nicotinic receptors. The total number of each of the toxin F binding sites was unaffected by culture conditions that are known to influence the extent to which these sympathetic neurons synthesize norepinephrine or acetylcholine. Autoradiographic analysis performed under saturating binding conditions (80 nM [125I]toxin F) revealed that the density of [125I]toxin F binding at synaptic membranes was about 5000 sites/micron 2, either in the absence of any competing ligand or in the presence of 2 microM alpha-bungarotoxin. In the presence of 2 microM unlabeled toxin F, there was no detectable binding at synapses. The density of these toxin F-specific sites was at least 80-fold higher at synaptic membranes than elsewhere. On the other hand, the data suggest that the toxin F binding site shared with alpha-bungarotoxin is exclusively extrasynaptic. Two micromolar alpha-bungarotoxin decreased the density of [125I]toxin F binding at non-synaptic sites by approximately two-thirds. These experiments support the hypothesis that toxin F blocks cholinergic transmission in cultures of sympathetic neurons by binding to nicotinic receptors and suggests that these receptors are highly clustered at synaptic membranes.

Animals↗
PubMed1988

Target specificity of neuropeptide Y-immunoreactive cranial parasympathetic neurons.

We recently showed that neuropeptide Y (NPY)-like immunoreactivity occurs in subpopulations of neurons in 3 cranial parasympathetic ganglia: the otic, sphenopalatine, and ciliary. The present work identifies the target tissues innervated by cranial parasympathetic NPY-immunoreactive neurons. Plexuses of NPY-immunoreactive fibers were observed in the parotid gland, the target of the otic ganglion, and in the intraorbital lacrimal gland and palate, targets of the sphenopalatine ganglion. NPY-immunoreactive fibers of apparent parasympathetic origin innervated glandular acini in all 3 structures and were also present around small blood vessels in the parotid and intraorbital lacrimal glands. These fibers were presumed to be parasympathetic because they were not affected by removal of the superior cervical ganglion and because their distribution was coextensive with that of vasoactive intestinal polypeptide (VIP) immunoreactivity, which we have previously shown to be colocalized with NPY in the cell bodies of otic and sphenopalatine ganglion neurons. In contrast, no NPY-immunoreactive fibers were observed in the iris or ciliary body of acutely sympathectomized rats, suggesting that NPY-immunoreactive neurons in the ciliary ganglion do not normally transport detectable levels of NPY to their terminals. The target specificities of cranial parasympathetic NPY-immunoreactive neurons are different from those of sympathetic NPY-immunoreactive neurons. Sympathetic NPY-immunoreactive fibers innervated the iris and ciliary body, and the blood vessels but not the parenchymal cells of all the glands examined. In contrast, parasympathetic NPY-immunoreactive fibers primarily innervated glandular acini. NPY-immunoreactive neurons in the sphenopalatine ganglion displayed an additional level of specificity in their projection pattern in that they innervated only a subset of the ganglion's array of target glands: they innervated the intraorbital lacrimal gland and the seromucous glands of the palate but not the exorbital lacrimal gland or the glands of the nasal mucosa. The finding that NPY immunoreactivity is present in the parasympathetic innervation of secretory acini in several craniofacial glands raises the possibility that NPY plays a role in the parasympathetic control of glandular secretion. The observed overlap in the distributions of NPY- and VIP-immunoreactive fibers in these glands further suggests that NPY may interact with VIP to stimulate secretion.

Animals↗
PubMed1987

Development and properties of the secretory response in rat sweat glands: relationship to the induction of cholinergic function in sweat gland innervation.

Previous studies suggest that the sympathetic innervation of the sweat glands in the rat is initially noradrenergic and during development undergoes a transition in neurotransmitter phenotype to become cholinergic. To characterize this system and its development further, we have examined the adrenergic and cholinergic components of the secretory response in adult and immature rats and have studied the onset of sweating in the plantar sweat glands of developing rats. Stimulation of the sciatic nerve in adult rats elicited a secretory response which was completely blocked by the cholinergic antagonist, atropine, and was unaffected by adrenergic antagonists, indicating that nerve-evoked secretion was cholinergic. In adult rats, the sweat glands were quite sensitive to cholinergic agonists. In addition to acetylcholine, the mature sweat gland innervation contains vasoactive intestinal peptide (VIP). In some rats, the injection of VIP alone elicited a secretory response which was blocked by atropine, suggesting that the response to VIP was mediated cholinergically. In contrast to cholinergic agonists, the glands responded relatively infrequently and with reduced volumes of sweat to the alpha- and beta-adrenergic agonists 6-fluoronorepinephrine and isoproterenol. However, when VIP, which is a potent vasodilator, was simultaneously injected with adrenergic agonists, glands in many of the injected footpads exhibited a secretory response. The response to adrenergic agonists in combination with VIP was reduced by atropine and by phentolamine plus propranolol, but was blocked completely only by a combination of the three antagonists, indicating that both adrenergic and cholinergic mechanisms were involved. In immature rats, sweating evoked by nerve stimulation first appeared at 14 days of age in 25% of the rats tested. Both the percentage of rats sweating and the number of active glands increased rapidly. At 16 days, 50% of the rats tested exhibited some active glands, and by 21 days all rats tested exhibited a secretory response. In 16-day-old rats, nerve-evoked sweating was almost completely inhibited by local injection of 1 microM atropine, but was unaffected by phentolamine and propranolol in concentrations up to 10 microM. Similarly, the glands were sensitive to 10 microM muscarine, but they exhibited no secretory response to the alpha-adrenergic agonists, clonidine and 6-fluoronorepinephrine, nor to the beta-adrenergic agonist, isoproterenol, at concentrations up to 50 microM. The simultaneous injection of VIP with adrenergic agonists did not reveal an adrenergically mediated secretory response in 16-day-old animals.(ABSTRACT TRUNCATED AT 400 WORDS)

Aging↗
PubMed1987

Neuropeptide Y-like immunoreactivity in rat cranial parasympathetic neurons: coexistence with vasoactive intestinal peptide and choline acetyltransferase.

Neuropeptide Y (NPY) is widely distributed in the sympathetic nervous system, where it is colocalized with norepinephrine. We report here that NPY-immunoreactive neurons are also abundant in three cranial parasympathetic ganglia, the otic, sphenopalatine, and ciliary, in the rat. High-performance liquid chromatographic analysis of the immunoreactive material present in the otic ganglion indicates that this material is very similar to porcine NPY and indistinguishable from the NPY-like immunoreactivity present in rat sympathetic neurons. These findings raise the possibility that NPY acts as a neuromodulator in the parasympathetic as well as the sympathetic nervous system. In contrast to what has been observed for sympathetic neurons, NPY-immunoreactive neurons in cranial parasympathetic ganglia do not contain detectable catecholamines or tyrosine hydroxylase (EC 1.14.16.2) immunoreactivity, and many do contain immunoreactivity for vasoactive intestinal peptide and/or choline acetyltransferase (EC 2.3.1.6). These findings suggest that there is no simple rule governing coexpression of NPY with norepinephrine, acetylcholine, or vasoactive intestinal peptide in autonomic neurons. Further, while functional studies have indicated that NPY exerts actions on the peripheral vasculature which are antagonistic to those of acetylcholine and vasoactive intestinal peptide, the present results raise the possibility that these three substances may have complementary effects on other target tissues.

Animals↗
PubMed1987

Catecholaminergic properties of cholinergic neurons and synapses in adult rat ciliary ganglion.

Parasympathetic neurons of the ciliary ganglion are innervated by preganglionic cholinergic neurons whose cell bodies lie in the brain stem; the ganglion cells in turn provide cholinergic innervation to the intrinsic muscles of the eye. Noradrenergic innervation of the iris is supplied by sympathetic neurons of the superior cervical ganglion. Using immunocytochemical and histochemical techniques, we have examined the ciliary ganglion of adult rats for the expression of cholinergic and noradrenergic properties. As expected, the postganglionic ciliary neurons possessed detectable levels of choline acetyltransferase immunoreactivity (ChAT-IR). Unexpectedly, many ciliary neurons also exhibited immunoreactivity for tyrosine hydroxylase (TH-IR). Some had dopamine beta-hydroxylase-like (DBH-IR) immunoreactivity, but none contained detectable catecholamines, even after treatment with nialamide and L-DOPA. A sparse plexus of fibers exhibiting faint TH-IR was present in the irises of acutely sympathectomized rats. The terminals of preganglionic axons in the ciliary ganglion exhibited not only immunoreactivity for ChAT, but also for TH and contained stores of endogenous catecholamine. Neither ciliary neurons nor their preganglionic innervation accumulated detectable stores of exogenous catecholamines. Rats sympathectomized as neonates by treatment with 6-hydroxydopamine subsequently had a greater proportion of neurons possessing detectable TH-IR in the ciliary ganglion; both the TH-IR perikarya and their axons in the iris were more intensely immunofluorescent. TH-IR was present in the ciliary neuron cell bodies of mouse, guinea pig, and ferret. These species, however, lacked detectable TH-IR or catecholamine stores in preganglionic terminals. These observations indicate that mature, functionally cholinergic neurons from 2 different embryonic origins, postganglionic ciliary neurons derived from the neural crest and preganglionic neurons derived from the neural tube, display several catecholaminergic properties.

Animals↗
PubMed1986

Coexistence of calcitonin gene-related peptide and vasoactive intestinal peptide in cholinergic sympathetic innervation of rat sweat glands.

Immunoreactivity for calcitonin gene-related peptide (CGRP) has been localized with indirect immunofluorescence techniques in the cholinergic sympathetic fibers that innervate eccrine sweat glands in the rat. This innervation also contains vasoactive intestinal peptide-like immunoreactivity (VIP-IR). A small proportion of principal neurons in stellate and lumbar sympathetic ganglia which provide innervation to the sweat glands contain detectable CGRP-immunoreactivity. The CGRP-IR neurons are immunoreactive for VIP; however, many VIP-IR neurons in these ganglia do not contain detectable levels of CGRP-IR.

Animals↗
PubMed1986

Absence of the alpha and gamma subunits of 7S nerve growth factor in denervated rodent iris: immunocytochemical studies.

Immunocytochemical studies were performed to determine if denervated rodent iris produces nerve growth factor (NGF) in a form chemically similar to that of the 7S NGF complex in mouse submandibular glands. Antisera to the alpha, beta, and gamma subunits of 7S NGF were raised in rabbits and characterized on immunoblots of SDS-containing polyacrylamide gels. Antisera were applied to stretch preparations of rat and mouse irides that were cultured for periods of 2 to 6 days or sympathetically denervated by superior cervical ganglionectomy and left in situ 4 days. Antibody binding was visualized by indirect immunofluorescence. In control studies done on plastic sections of mouse submandibular glands, antisera co-localized the three subunits of 7S NGF within secretory granules of granular tubule cells. In denervated rat iris, beta NGF immunoreactivity was evident in a cellular plexus that resembled in distribution and morphology nerve fibers in the normal iris, in agreement with a previous study (R.A. Rush (1984). Nature (London) 312, 364-367). Identical staining patterns were observed in mouse iris. In neither rat or mouse, however, did the nerve-like processes stain with antibodies suggests that the NGF-like protein in denervated rodent iris is not synthesized as part of the 7S NGF complex. Iris also did not react with antibodies to epidermal growth factor, a protein co-localized with NGF in mouse submandibular glands and in guinea pig prostate.

Animals↗
PubMed1986

Vasoactive intestinal peptide: vasodilatation and cyclic AMP generation.

Vasoactive intestinal peptide (VIP) is a putative neurotransmitter that causes vasodilation when injected intravenously. To learn more about the vasodilator actions of VIP, studies were performed on the rat mesenteric arterial bed and on cultured smooth muscle cells derived from both the rat mesenteric artery and aorta. In isolated perfused rat mesenteric artery beds, VIP caused relaxation at the threshold concentration of less than 1 nM and a maximal fall in perfusion pressure similar to that obtained with isoproterenol. VIP stimulated adenylate cyclase in cultured vascular smooth muscle cells in a concentration range of 10(-10)-10(-6) M; maximum activity was increased 2.7 +/- 0.6- and 3.4 +/- 0.4-fold above basal in mesenteric and aortic smooth muscle cells, respectively. The threshold and the half-maximal concentrations of VIP for adenylate cyclase stimulation were more than 50-fold lower than those for both prostaglandin E1 and isoproterenol in cultured mesenteric and aortic cells. Similar effects on cyclic AMP generation in response to VIP were observed in isolated rat mesenteric artery rings. Immunocytochemical examination of the rat mesenteric artery bed in situ demonstrated dense innervation of small- and medium-size vessels with nerve fibers containing VIP-like immunoreactivity. Thus VIP is present in the rat mesenteric artery, a peripheral arterial bed, and is a potent vasodilator that can activate vascular smooth muscle adenylate cyclase and thus potentially contribute to the regulation of vascular tone.

Adenylyl Cyclases↗
PubMed1986

Synaptic functions in rat sympathetic neurons in microcultures. II. Adrenergic/cholinergic dual status and plasticity.

This is the second in a series of papers that describes the use of a sensitive microculture procedure to investigate the transmitter status of sympathetic neurons. Cultured immature principal neurons, dissociated from the superior cervical ganglia of newborn rats, are known to be plastic with respect to transmitter status; under certain culture conditions, populations of neurons that display (at least) adrenergic properties at the outset can be induced to display a variety of cholinergic properties, including the formation of functional neuron-neuron cholinergic synapses, as adrenergic properties decline. With the microculture procedure described in the preceding paper (Furshpan et al., 1986a), we have examined the transmitter status of individual neonate-derived neurons during this transition. Many such neurons secreted both norepinephrine and ACh (adrenergic/cholinergic dual function); examination of such neurons with the EM revealed a mixed population of synaptic vesicles. Direct evidence for a transition via this dual status was obtained by serial physiological assays of 14 neurons. The neonate-derived neurons were markedly heterogeneous in the rate of change of transmitter status. Principal neurons derived from adult superior cervical ganglia also displayed dual status, but the incidence was lower than in neonate-derived neurons cultured for similar periods. In preliminary serial assays of adult-derived neurons, many of the neurons did not acquire detectable cholinergic function, but in two cases evidence consistent with plasticity was obtained. While it is known that several types of neurons will form functional junctions in the presence of agents that block electrical activity, sympathetic principal neurons have apparently not been tested. In microculture, neuron-neuron synapses and junctions with cardiac myocytes were formed by sympathetic neurons grown chronically in the presence of blocking concentrations of TTX and hexamethonium.

Acetylcholine↗
PubMed1986

Synaptic functions in rat sympathetic neurons in microcultures. I. Secretion of norepinephrine and acetylcholine.

This is the first of a series of four papers that describes the use of a sensitive "microculture" procedure for examining the neurotransmitter profile of a neuron by assaying the transmitter(s) it releases. Sympathetic principal neurons isolated from the superior cervical ganglia of neonatal or adult rats were grown for 10 d to several months on small islands of cardiac myocytes (island diameter, ca. 0.5 mm). To assay transmitter status a neuron and a myocyte in the same microculture were impaled with microelectrodes, the neuron was stimulated and the pharmacology of the effect(s) on the group of electrically coupled myocytes, and on the neuron itself, was investigated. Because the growing axonal processes were confined to the island, the innervation of the myocytes became dense; transmission from neuron to myocytes occurred reliably and was often intense. Most experiments were done on islands containing only a single neuron so that the observed effect(s) on the myocytes could be confidently assigned to that neuron. After the physiological assay, the fine structure or cytochemistry of the neuron was often examined. With single-neuron microcultures the physiology and anatomy of the neuron, including the fine structure of its synaptic endings and varicosities, could be correlated unambiguously. During the course of this work, we have observed five pharmacologically distinct effects exerted on the myocytes by either neonate- or adult-derived neurons. Three of these effects, one exerted at least in large part by adenosine and the others by agents still under study (one appears to be 5-HT), are described by Furshpan et al. (1986), Matsumoto et al. (in press), and D. Sah and S. G. Matsumoto (unpublished observations). This paper is concerned with evidence for secretion by these neurons of norepinephrine (NE) and acetylcholine (ACh). The physiological effects of the secretion of these two substances onto the myocytes (excitation and inhibition, respectively) were generally similar to those reported in vivo. The minimal latencies of the responses were short, probably due to the high density of innervation. ACh secreted by a neuron onto itself, at autapses, evoked fast nicotinic EPSPs. We have not detected autaptic effects attributable to the secretion of NE. A minority of the neurons were detectably only adrenergic or only cholinergic. The incidence of these transmitter states was strongly dependent on culture age and culture conditions; in a heterogeneous group of about 300 reasonably well-characterized neurons about 17% (12% of neonate-derived) were apparently purely adrenergic and about 10% (13% of neonate-derived) were apparently purely cholinergic.(ABSTRACT TRUNCATED AT 400 WORDS)

Acetylcholine↗
PubMed1985

Neonatal treatment with nerve growth factor antiserum eliminates cholinergic sympathetic innervation of rat sweat glands.

Most mammalian sympathetic neurons are noradrenergic, and their dependence upon nerve growth factor (NGF) for survival during development is well established. A minor population of sympathetic neurons, including those that innervate sweat glands, is cholinergic. To determine whether cholinergic sympathetic neurons, like their noradrenergic counterparts, require NGF during development, neonatal rats were treated with NGF-antiserum and 3 weeks later their sweat glands were examined for the presence of innervation. Acetylcholinesterase (AChE) staining and vasoactive intestinal polypeptide-like immunoreactivity (VIP-IR) which mark the mature sweat gland innervation were absent from the sweat glands of the anti-NGF treated animals. Further, when the glands were examined with the electron microscope, no axons or nerve terminals were evident. These observations indicate that the elaboration of the sweat gland plexus is NGF-dependent and suggest that at least one population of cholinergic sympathetic neurons in the rat requires NGF for survival. Our findings are consistent with the idea that during development NGF is a required trophic factor not only for noradrenergic sympathetic but also for cholinergic sympathetic neurons.

Acetylcholinesterase↗