Ketamine: evidence of tissue specific inhibition of neuronal and extraneuronal catecholamine uptake processes

Ketamine (1.1 X 10(-5) to 3.7 X 10(-4) M) potentiated catecholamine responses of rat anococcygeus muscle and rabbit aorta in vitro. In the anococcygeus, potentiation was abolished by cocaine (2.9 X 10(-5) M) pretreatment or by chemical sympathectomy using 6-hydroxydopamine (6-OHDA), but was unaffected by pretreatment with the extraneuronal uptake inhibitor cortisol (8.3 X 10(-5) M), or the catechol-O-methyltransferase inhibitor tropolone (2.4 X 10(-4) M). The action of ketamine mimicked the potentiating effect of cocaine on tyramine responses. In contrast, the potentiation by ketamine in rabbit aorta was unaffected by cocaine or 6-OHDA but was abolished by cortisol or tropolone; and ketamine potentiated tyramine responses, whereas cocaine inhibited them. Thus the mechanism of action by which ketamine produces potentiation of catecholamines in these two tissues is completely different. These results suggest that ketamine has the unusual ability to block neuronal and extraneuronal uptake and that the predominating mechanism will depend on the type of tissue examined and the morphology of its adrenergic innervation.     

Adrenomedullary Secretion of DOPA, Catecholamines, Catechol Metabolites, and Neuropeptides

Abstract: Catecholamines and their metabolites have been proposed as markers of sympathetic nervous system stimulation. However, the adrenal medulla is a rich source of catecholamines and catecholamine metabolites and may play a significant role in plasma levels of these compounds. In addition to adrenal catecholamine metabolite efflux, the role of the catecholamine precursor 3,4-dihydroxyphenylalanine (DOPA) has not been fully evaluated. The simultaneous effluxes of catecholamines, metabolites, DOPA, and neuropeptides were measured in perfusates from isolated dog adrenals. The relative abundance of compounds detected consistently during unstimulated conditions was epinephrine ≫ norepinephrine > 3,4-dihydroxyphenylglycol > metanephrine > normetanephrine > dopamine > 3,4-dihydroxyphenylacetic acid > 3-methoxy-4-hydroxyphenylglycol ≥ DOPA ≫ [Met]enkephalin ≫ neuropeptide Y. Effluxes of analytes were not affected by cocaine and the ratios of catecholamines to metabolites increased dramatically with carbachol stimulation, consistent with negligible reuptake into adrenal cells. Thus, most of the 3,4-dihydroxyphenylglycol is expected to be derived from epinephrine and norepinephrine subsequent to translocation from chromaffin vesicles into the cytosol. The efflux of DOPA increased dramatically during stimulation with 30 µ M carbachol in a calcium-dependent manner. Efflux of DOPA during the initial stabilization period of the perfusion preparation declined exponentially, in parallel with the effluxes of the catecholamines and neuropeptides but not with metabolites. Evoked release of DOPA was Ca²⁺-dependent. These data suggest that DOPA can be stored and released exocytotically from chromaffin granules.     

Tissue specific regulation of "peripheral-type" benzodiazepine receptor density after chemical sympathectomy

The characteristics of [3H]Ro 5-4864 binding to "peripheral" benzodiazepine receptors (PBR) in the central nervous system and peripheral tissues were examined after chemical sympathectomy with 6-hydroxydopamine (6-OHDA). One week after the intracisternal administration of 6-OHDA, the number of [3H]Ro 5-4864 binding sites (Bmax) in the hypothalamus and striatum increased 41 and 50%, respectively, concurrent with significant reductions in catecholamine content. An increase (34%) in the Bmax of [3H]Ro 5-4864 to cardiac ventricle was observed one week after parenteral 6-OHDA administration. In contrast, the Bmax of [3H]Ro 5-4864 to pineal gland decreased 48% after 6-OHDA induced reduction in norepinephrine content. The Bmax values for [3H]Ro 5-4864 binding to other tissues (including lung, kidney, spleen, cerebral cortex, cerebellum, hippocampus and olfactory bulbs) were unaffected by 6-OHDA administration. The density of pineal, but not cardiac PBR was also reduced after reserpine treatment, an effect reversed by isoproterenol administration. These findings demonstrate that alterations in sympathetic input may regulate the density of PBR in both the central nervous system and periphery in a tissue specific fashion.     

Role of the sympathetic innervation on uterine electromyographic activity in nonpregnant ovariectomized sheep under estrogen supplementation.

The aims of the present study were to characterize the sympathetic innervation of the nonpregnant sheep uterus, to determine the catecholamine content in myometrium (MYO) and endometrium, and to study the effects of chemical sympathectomy (CHSPX) on uterine catecholamine content and on uterine electromyographic (EMG) activity recorded from the MYO and mesometrium (MESO) in the nonpregnant ovariectomized sheep. After synchronization of estrus, 9 nonpregnant sheep were anesthetized with halothane, ovariectomized, and fitted with vascular catheters and EMG electrodes. Estradiol-17 beta was administered intravascularly at a rate of 50 micrograms/24 h for 10 days. CHSPX was induced with 6-hydroxy dopamine (20 mg/kg). Uterine tissues were obtained for determination of catecholamine content by HPLC and for immunocytochemical staining using an antibody against tyrosine hydroxylase (TH). In nonpregnant ovariectomized sheep, TH immunostaining was present in nerve fibers located in endometrium and MYO. In all layers of the uterus, catecholamine fibers were found in the proximity of blood vessels as well as in defined regions of the parenchyma. Throughout the uterus, norepinephrine content and TH immunostaining were dramatically decreased after CHSPX. CHSPX decreased uterine short EMG event activity in both MYO and MESO. Contracture-type activity was not affected in MYO and was increased in MESO. We conclude that sympathetic innervation modulates the MYO and MESO EMG activity in nonpregnant ovariectomized sheep under estradiol supplementation, and that the removal of the sympathetic innervation induces a decrease in the spontaneous activity.     

Adipocytes as a new source of catecholamine production

Catecholamines are an important regulator of lipolysis in adipose tissue. Here we show that rat adipocytes, isolated from mesenteric adipose tissue, express genes of catecholamine biosynthetic enzymes and produce catecholamines de novo. Administration of tyrosine hydroxylase inhibitor, alpha-methyl-p-tyrosine, in vitro significantly reduced concentration of catecholamines in isolated adipocytes. We hypothesize that the sympathetic innervation of adipose tissues is not the only source of catecholamines, since adipocytes also have the capacity to produce both norepinephrine and epinephrine.     

Chronic depolarization stimulates norepinephrine transporter expression via catecholamines

Chronic depolarization increases norepinephrine (NE) uptake and expression of the norepinephrine transporter (NET) in sympathetic neurons, but the mechanisms are unknown. Depolarization of sympathetic neurons stimulates catecholamine synthesis, and several studies suggest that NET can be regulated by catecholamines. It is not clear if the depolarization-induced increase in NET is because of nerve activity per se, or is secondary to elevated catecholamines. To determine if induction of NET mRNA was a result of increased catecholamines, we used pharmacological manipulations to (i) inhibit tyrosine hydroxylase activity in neurons depolarized with 30 mm KCl, thereby preventing increased catecholamines, or (ii) stimulate tyrosine hydroxylase activity in the absence of depolarization. Inhibiting the depolarization-induced increase in catecholamines prevented the up-regulation of NET mRNA, but did not block the increase in tyrosine hydroxylase (TH) mRNA. Furthermore, stimulating catecholamine production in the absence of depolarization elevated NE uptake, NET protein, and NET mRNA in sympathetic neurons. Similarly, elevating endogenous catecholamines in SK-N-BE2M17 neuroblastoma cells increased NE uptake and NET expression. These data suggest that chronic depolarization of sympathetic neurons induces NET expression through increasing catecholamines, and that M17 neuroblastoma cells provide a model system in which to investigate catechol regulation of NET expression.     

Effect of dietary proteins and carbohydrates on urinary and sympathoadrenal catecholamines

We previously observed that administration of tyrosine to rats or humans elevated urinary dopamine, norepinephrine and epinephrine levels. The present studies examine the effects on these urinary catecholamines of varying the ratio of protein to carbohydrate in the diets.Rats consumed diets containing 0, 18 or 40% protein (76, 58 and 36% carbohydrate respectively) for 8 days. The stress of consuming the protein-free food was associated with a 16% weight reduction, and with significantly lower serum, heart and brain tyrosine levels than those noted in rats eating the 18 or 40% protein diets. Absence of protein from the diet also decreased urinary levels of dopamine and DOPA but increased urinary norepinephrine and epinephrine, probably by increasing sympathoadrenal discharge; it also increased the excretion of DOPA in animals pretreated with carbidopa, a DOPA decarboxylase inhibitor. Carbidopa administration decreased urinary dopamine, norepinephrine and epinephrine as expected; however, among carbidopa-treated rats urinary norepinephrine and epinephrine concentrations were highest for animals consuming the protein-free diet, again suggesting enhanced release of stored catecholamines from sympathoadrenal cells. The changes in urinary catecholamines observed in animals eating the protein-free diet were similar to those seen in rats fasted for 5 days: dopamine levels fell sharply while norepinephrine and epinephrine increased.These data indicate that the effects of varying dietary protein and carbohydrate contents on dopamine secretion from peripheral structures differ from its effects on structures secreting the other two catecholamines. Protein consumption increases dopamine synthesis and release probably by making more of its precursor, tyrosine, available to peripheral dopamine-producing cells; it decreases urinary norepinephrine and epinephrine compared with that seen in protein-deprived animals, probably by diminishing the firing of sympathetic neurons and adrenal chromaffin cells.     

Expression of neuronal markers suggests heterogeneity of chick sympathoadrenal cells prior to invasion of the adrenal anlagen

We have analyzed the distribution of neural crest-derived precursors and the expression of catecholaminergic and neuronal markers in developing adrenal tissue of chick embryos. Undifferentiated neural crest cells are found in presumptive adrenal regions from embryonic day 3 (E3) onward. An increasing proportion of cells expressing tyrosine hydroxylase (TH) mRNA indicates catecholaminergic differentiation of precursors not only in primary sympathetic ganglia, but also in presumptive adrenal regions. Whereas precursors and differentiating cells show mesenchymal distribution until E5, discrete adrenal anlagen form during E6. Even during E5, catecholaminergic cells with low or undetectable neurofilament M (NF-M) mRNA expression prevail in positions at which adrenal anlagen become distinct during E6. The predominance of TH-positive and NF-M-negative cells is maintained throughout embryogenesis in adrenal tissue. RNA encoding SCG10, a pan-neuronal marker like NF-M, is strongly expressed throughout adrenal anlagen during E6 but is found at reduced levels in chromaffin cells compared with neuronal cells at E15. Two additional neuronal markers, synaptotagmin 1 and neurexin 1, are expressed at low to undetectable levels in developing chromaffin cells throughout embryogenesis. The developmental regulation of neuronal markers shows at least three different patterns among the four mRNAs analyzed. Importantly, there is no generalized downregulation of neuronal markers in developing adrenal anlagen. Thus, our observations question the classical concept of chromaffin differentiation from a common sympathoadrenal progenitor expressing neuronal properties and suggest alternative models with changing instructive signals or separate progenitor populations for sympathetic neuronal and chromaffin endocrine cells.Chaya Kalcheim and Klaus Unsicker are supported by the Deutsche Forschungsgemeinschaft (SFB 488)     

Dopamine and norepinephrine in the alimentary tract changes after chemical sympathectomy and surgical vagotomy

The aim of this study was to examine the distribution of dopamine and norepinephrine in the proximal alimentary tract of the rat and to assess the contributions of sympathetic and vagal fibers to the tissue concentrations of both catecholamines. Tissues were extracted in perchloric acid and the catecholamines were separated by high pressure liquid chromatography and detected electrochemically. In untreated rats (controls) both catecholamines were concentrated in the gastric muscle but norepinephrine levels were 6-8 times higher (corpus, dopamine 35 +/- 7 ng . g-1, norepinephrine 265 +/- 50 ng . g-1, mean +/- SE, n = 6). In the mucosa norepinephrine concentrations were 10-12 times higher (corpus, dopamine 12 +/- 3 ng . g-1, norepinephrine 140 +/- 26 ng . g-1). Chemical sympathectomy (6 hydroxydopamine, 100 mg . kg-1 ip 3 days) significantly reduced dopamine concentrations in muscle and norepinephrine in muscle, mucosa, pylorus and duodenum. In all tissues the effects on norepinephrine were greater. Surgical vagotomy significantly reduced dopamine concentrations in the gastric muscle, but not the mucosa. Norepinephrine concentrations in the stomach of vagotomized rats were significantly reduced only in the pylorus. Differences in the relative concentrations of dopamine and norepinephrine in gastric tissues of the normal rat and differences in the effects of sympathectomy and vagotomy suggest that dopamine and norepinephrine exist, to an extent, in separate populations of cells and that dopamine is not merely a precursor of norepinephrine. Gastric mucosal dopamine, which was mainly unaffected by either treatment, may exist in APUD cells.     

Central catecholamine depletion inhibits peripheral lymphocyte responsiveness in spleen and blood

Experimental and clinical evidence has demonstrated extensive communication between the CNS and the immune system. To analyse the role of central catecholamines in modulating peripheral immune functions, we injected the neurotoxin 6-hydroxydopamine (6-OHDA) i.c.v. in rats. This treatment significantly reduced brain catecholamine content 2, 4 and 7 days after injection, and in the periphery splenic catecholamine levels were reduced 4 days after treatment. Central catecholamine depletion induced an inhibition of splenic and blood lymphocyte proliferation and splenic cytokine production and expression (interleukin-2 and interferon-gamma) 7 days after injection. In addition, central treatment with 6-OHDA reduced the percentage of spleen and peripheral blood natural killer (CD161 +) cells, and T-cytotoxic (CD8 +) cells in peripheral blood. The reduction in splenocyte proliferation was not associated with a glucocorticoid alteration but was completely abolished by prior peripheral sympathectomy. These data demonstrate a crucial role of central and peripheral catecholamines in modulating immune function.     

Neuroendocrine properties of intrinsic cardiac adrenergic cells in fetal rat heart

Intrinsic cardiac adrenergic (ICA) cells in developing rat heart constitute a novel adrenergic signaling system involved in cardiac regulation. Regulatory mechanisms of ICA cells remain to be defined. Immunohistochemical study of fetal rat hearts demonstrated ICA cells with catecholamine biosynthetic enzyme tyrosine hydroxylase (TH) and phenylethanolamine N-methyltransferase (PNMT). The mRNA of TH and PNMP was also detected in fetal rat hearts before sympathetic innervation. Immunoreactivity of norepinephrine transporter (NET) was localized to ICA cells in rat heart tissue and primary cell culture. For the functional study, the activity of intracellular Ca2+ concentration ([Ca2+]i) transients was quantified by a ratio fluorescent spectrometer in cultured ICA cells and myocytes. ICA cells generated spontaneous [Ca2+]i transients that were eliminated by tetrodotoxin or Ca(2+)-free solutions and showed greatly reduced amplitude with the addition of L-type Ca2+ channel blocker nifedipine. [3H]norepinephrine studies demonstrate release and uptake of norepinephrine. Functional interaction between catecholamines produced by the ICA cells and cocultured myocytes was evident by the effect of the beta-adrenergic blocker atenolol eliciting a dose-dependent reduction in the amplitude and frequency of [Ca2+]i transients of beating myocytes. Hypoxia inhibited [Ca2+]i transient activity of ICA cells, which subsequently produced a reoxygenation-mediated rebound augmentation of [Ca2+]i transients. We conclude that ICA cells are capable of catecholamine synthesis, release, and uptake. They generate spontaneous [Ca2+]i transient activity that can be regulated by oxygen tension. ICA cells may provide an alternative adrenergic supply to maintain cardiac contractile and pacemaker function at rest and during stress in the absence of sympathetic innervation.     

Autoimmune sympathectomy in fetal rabbits

An autoimmune model for in utero immunosympathectomy of fetal rabbits was developed. Non-pregnant, female rabbits were injected with purified nerve growth factor and then bred after confirmation of high titers of anti-nerve growth factor antiserum. Fetuses were delivered and sacrificed at 27 and 31 days gestation and tissue norepinephrine concentration was used as an index of sympathetic innervation. There were significant reductions in tissue norepinephrine at both gestational ages. At 31 days there was a 32% reduction in lung norepinephrine concentration, 46% in the heart and 60% in brown adipose tissue. Corresponding reductions at 27 days were 68% for lung, 44% for heart and 49% for brown adipose tissue. Adrenal catecholamine content was unaffected but para-aortic gland catecholamines were slightly increased. Pulmonary beta adrenergic receptors showed a 30% up regulation in response to dennervation. Carcass weight was reduced 15% to 11% in the dennervated animals. These results demonstrate that dependence of organ sympathetic innervation on nerve growth factor can be demonstrated as early as 27 days gestation. This is a useful model to study the timing and dependence of organ sympathetic innervation on nerve growth factor and the factors which regulate this dependence.     

Effects of natriuretic peptides (ANP, BNP, CNP) on catecholamine synthesis and TH mRNA levels in PC12 cells

Atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP) and C-type natriuretic peptide (CNP) are present in adrenal chromaffin cells, and are co-secreted with catecholamines suggesting that these natriuretic peptides (NPs) may modulate functions of chromaffin cells in an autocrine and/or paracrine manner. Therefore, we investigated the effects of NPs on tyrosine hydroxylase (TH: a rate-limiting enzyme in biosynthesis of catecholamine) mRNA in rat pheochromocytoma PC12 cells. It was also determined whether the cyclic GMP/cGMP-dependent protein kinase (cGMP/PKG) pathway was involved in theses effects. Finally, we examined the effects of NPs on intracellular catecholamine content to confirm increase of catecholamine synthesis following TH mRNA induction. NPs (0.1 microM) induced significant increases of the TH mRNA (ANP= BNP> CNP). Also, the effects of NPs on TH mRNA were mimicked by 8-bromo cyclic GMP (1mM), and were blocked by KT5823 (1 microM) (inhibitor PKG) or LY83583 (1 microM) (guanylate cyclase inhibitor). Moreover, NPs were shown to induce significant increases of intracellular catecholamine contents (ANP= BNP> CNP). These findings suggest that NPs induced increases of TH mRNA through cGMP/PKG dependent mechanisms, which, in turn, resulted in stimulation of catecholamine synthesis in PC12 cells.     

White adipose tissue lacks significant vagal innervation and immunohistochemical evidence of parasympathetic innervation

Converging evidence indicates that white adipose tissue (WAT) is innervated by the sympathetic nervous system (SNS) based on immunohistochemical labeling of a SNS marker (tyrosine hydroxylase [TH]), tract tracing of WAT sympathetic postganglionic innervation, pseudorabies virus (PRV) transneuronal labeling of WAT SNS outflow neurons, and functional evidence from denervation studies. Recently, WAT para-SNS (PSNS) innervation was suggested because local surgical WAT sympathectomy (sparing hypothesized parasympathetic innervation) followed by PRV injection yielded infected cells in the vagal dorsomotor nucleus (DMV), a traditionally-recognized PSNS brain stem site. In addition, local surgical PSNS WAT denervation triggered WAT catabolic responses. We tested histologically whether WAT was parasympathetically innervated by searching for PSNS markers in rat, and normal (C57BL) and obese (ob/ob) mouse WAT. Vesicular acetylcholine transporter, vasoactive intestinal peptide and neuronal nitric oxide synthase immunoreactivities were absent in WAT pads (retroperitoneal, epididymal, inguinal subcutaneous) from all animals. Nearly all nerves innervating WAT vasculature and parenchyma that were labeled with protein gene product 9.5 (PGP9.5; pan-nerve marker) also contained TH, attesting to pervasive SNS innervation. When Siberian hamster inguinal WAT was sympathetically denervated via local injections of catecholaminergic toxin 6-hydroxydopamine (sparing putative parasympathetic nerves), subsequent PRV injection resulted in no central nervous system (CNS) or sympathetic chain infections suggesting no PSNS innervation. By contrast, vehicle-injected WAT subsequently inoculated with PRV had typical CNS/sympathetic chain viral infection patterns. Collectively, these data indicate no parasympathetic nerve markers in WAT of several species, with sparse DMV innervation and question the claim of PSNS WAT innervation as well as its functional significance.     

Catecholaminergic control of plasma growth hormone and thyrotropin levels in hypophysectomized rats bearing ectopic pituitary transplants

Transplantation of the anterior pituitary to an ectopic site leads to stimulation of PRL secretion and suppression of the release of other adenohypophyseal hormones. We have previously reported that precursors and blockers of catecholamine synthesis can affect PRL release from the ectopic pituitary. In the present study we have measured the effects of L-3,4-dihydroxyphenylalanine (DOPA), DL-threo-dihydroxyphenylserine (DOPS), alpha-methyl-para-tyrosine (alpha-mpt) and diethyldithiocarbamate (ddc) on plasma growth hormone (GH) and thyrotropin (TSH) levels in hypophysectomized rats with pituitary transplants under the renal capsule. In these animals, peripheral plasma GH levels were elevated by a precursor (DOPA) and reduced by a blocker (alpha-mpt) of catecholamine synthesis. Plasma TSH levels were increased by a precursor (DOPS) and reduced by a blocker (ddc) of norepinephrine synthesis. We suspect that GH and TSH present in the circulation of pituitary-grafted animals were derived, in part, from the ectopic pituitary tissue and suggest that the small but detectable secretion of hormones other than PRL in this animal model is under the control of endogenous catecholamines.     

Neurofilament immunoreactivity and acetylcholinesterase activity in the developing sympathetic tissues of the rat

Summary In this study, the ontogenetic appearance of three neuronal markers, tyrosine hydroxylase (TH), neurofilament (NF) proteins and acetylcholinesterase (AChE), have been compared in the neural tube and derivatives of the neural crest with special consideration on developing rat sympathetic tissues. The tree markers appeared for the first time on embryonic day E 12.5. At this age, NF immunoreactivity was located in the cells on the ventro- and dorsolateral edges of the neural tube, i.e., in the regions where the cells had reached the postmitotic stage. In addition, on day E 12.5, NF-immunoreactive fibers were located in the dorsal and ventral roots and the spinal and sympathetic ganglia. This suggests rapid extension of neurites. In contrast to NF, AChE first appeared on day E 12.5 in cell somata of spinal and sympathetic ganglia ond only after that in axons. Thus, it can be considered as a marker of differentiating neuronal cell bodies. In the developing sympathoadrenal cells, TH is expressed before NF and AChE. However, the migrating TH immunoreactive sympathetic cells are constantly followed by NF immunoreactive fibers, suggesting that sympathetic tissues may receive innervation from preganglionic axons at the very beginning of their ontogeny. During the later development, all sympathetic tissues contain two major cell groups: 1) one with a moderate TH immunoreactivity, NF immunoreactivity and AChE activity and 2) the other with an intense TH immunoreactivity but lacking NF immunoreactivity or AChE activity. The former includes principal neurons, neuron-like cells of the paraganglia and noradrenaline cells of the adrenal medullae, and the latter includes ganglionic small intensely fluorescent (SIF) cells, paraganglionic cells and medullary adrenaline cells.     

Neurofilament immunoreactivity and acetylcholinesterase activity in the developing sympathetic tissues of the rat.

In this study, the ontogenetic appearance of three neuronal markers, tyrosine hydroxylase (TH), neurofilament (NF) proteins and acetylcholinesterase (AChE), have been compared in the neural tube and derivatives of the neural crest with special consideration on developing rat sympathetic tissues. The tree markers appeared for the first time on embryonic day E 12.5. At this age, NF immunoreactivity was located in the cells on the ventro- and dorsolateral edges of the neural tube, i.e., in the regions where the cells had reached the postmitotic stage. In addition, on day E 12.5, NF-immunoreactive fibers were located in the dorsal and ventral roots and the spinal and sympathetic ganglia. This suggests rapid extension of neurites. In contrast to NF, AChE first appeared on day E 12.5 in cell somata of spinal and sympathetic ganglia and only after that in axons. Thus, it can be considered as a marker of differentiating neuronal cell bodies. In the developing sympathoadrenal cells, TH is expressed before NF and AChE. However, the migrating TH immunoreactive sympathetic cells are constantly followed by NF immunoreactive fibers, suggesting that sympathetic tissues may receive innervation from preganglionic axons at the very beginning of their ontogeny. During the later development, all sympathetic tissues contain two major cell groups: 1) one with a moderate TH immunoreactivity, NF immunoreactivity and AChE activity and 2) the other with an intense TH immunoreactivity but lacking NF immunoreactivity or AChE activity. The former includes principal neurons, neuron-like cells of the paraganglia and noradrenaline cells of the adrenal medullae, and the latter includes ganglionic small intensely fluorescent (SIF) cells, paraganglionic cells and medullary adrenaline cells.     

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)     

Influence of alcohol on peripheral neurotransmitter function

Presently available evidence for the action of ethanol on the peripheral noradrenergic system is reviewed. Acute ethanol treatment produces an initial elevation in catecholamine levels in blood, and in the excretion of catecholamines, which may be followed by a decrease in catecholamine release. Correspondingly, a hyposensitivity of peripheral beta receptors has been noted. Other tissues such as vascular smooth muscles show a biphasic response to ethanol: a hypersensitivity to norepinephrine at low concentrations of ethanol, and hyposensitivity to norepinephrine at high ethanol concentrations. In the adrenal medulla the activity of various enzymes involved in catecholamine synthesis is increased by ethanol treatment. Chronic treatment with ethanol does not alter endogenous norepinephrine levels in heart, but does increase its turnover. Chronic treatment also lowers the number of beta-adrenergic receptors in the heart, and decreases the response to peripherally administered isoproterenol, which suggests a hyposensitivity of peripheral beta receptors. Tolerance may develop to the chronic effects of ethanol in the heart and the vas deferens, but in the adrenals the activity of enzymes for catecholamine synthesis remains elevated with chronic treatment. During withdrawal, there is generalized hyperexcitability of the sympathetic nervous system. Norepinephrine and amine turnover in the heart is increased, as is the activity of adrenal medullary enzymes involved in catecholamine synthesis.     

Leptin directly stimulates catecholamine secretion and synthesis in cultured porcine adrenal medullary chromaffin cells.

Leptin, a protein encoded by the ob gene, is an adipose tissue-derived signaling factor involved in body weight homeostasis. The hypothalamus is a major site of central action for leptin. However, mounting evidence indicates expression of leptin receptor mRNA in various peripheral organs including the adrenal medulla. Therefore, we investigated the effects of leptin on catecholamine secretion and synthesis in cultured porcine adrenal medullary chromaffin cells. We initially confirmed the expression of leptin receptor (Ob-Rb) mRNA in cultured porcine adrenal medullary cells. Murine recombinant leptin (>==50 nM) strongly induced the release of both epinephrine (E) and norepinephrine (NE) from chromaffin cells. Removal of external Ca(2+) significantly suppressed these effects. Also, leptin (>==1 nM) enhanced nicotine-induced increases in E- and NE. Leptin (1, 10, 100 nM) significantly increased tyrosine hydroxylase (TH) (a rate-limiting enzyme in the biosynthesis of catecholamine) mRNA levels in a concentration-dependent manner. Furthermore, leptin (1, 10, 100 nM) significantly induced increases in cAMP levels, suggesting that the stimulatory effects on TH mRNA are mediated, at least in part, by the cAMP/protein kinase A pathway. These results indicate that leptin directly stimulates catecholamine release and synthesis, which in turn may potentiate the anti-obesity effects of leptin.