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B G Jenks

Publications and source records attributed to B G Jenks.

At least 37 records · Page 2Linked to original sources
PubMed1999

Serotonergic innervation of the pituitary pars intermedia of xenopus laevis.

At this point three brain centres are thought to be involved in the regulation of the melanotrope cells of the pituitary pars intermedia of Xenopus laevis: the magnocellular nucleus, the suprachiasmatic nucleus and the locus coeruleus. This study aims to investigate the existence of a fourth, serotonergic, centre controlling the melanotrope cells. In-vitro superfusion studies show that serotonin has a dose-dependent stimulatory effect on peptide release (1.6 x basal level at 10(-6) M serotonin) from single melanotrope cells. Retrograde neuronal tract tracing experiments, with the membrane probe FAST Dil applied to the pars intermedia, reveals retrogradely labelled neurones in the magnocellular nucleus, the suprachiasmatic nucleus, the locus coeruleus and the raphe nucleus. Of these brain centres, after immunocytochemistry only the raphe nucleus revealed serotonin-immunoreactive cell bodies. In addition, serotonin-immunoreactive cell bodies were found in the nucleus of the paraventricular organ, the posteroventral tegmental nucleus and the reticular istmic nucleus. In the pituitary, the pars nervosa, pars intermedia and pars distalis all reveal serotonin-immunoreactive nerve fibres. With immunocytochemical double-labelling for tyrosine hydroxylase and serotonin no colocalization of serotonin and tyrosine hydroxylase was observed in cell bodies in the brain, and in the pituitary hardly any colocalization was found in the nerve fibres. However, after in-vitro loading of neurointermediate lobes with serotonin, tyrosine hydroxylase and serotonin appear to coexist in a fibre network in the pars intermedia. On the basis of these data we propose that the melanotrope cells in the Xenopus pars intermedia are innervated by a 5-HT network originating in the raphe nucleus; this network represents the first identified stimulatory input to the pars intermedia of this species.

Animals↗
PubMed1998

Action currents generate stepwise intracellular Ca2+ patterns in a neuroendocrine cell.

It is believed that specific patterns of changes in the cytosolic-free calcium concentration ([Ca2+]i) are used to control cellular processes such as gene transcription, cell proliferation, differentiation, and secretion. We recently showed that the Ca2+ oscillations in the neuroendocrine melanotrope cells of Xenopus laevis are built up by a number of discrete Ca2+ rises, the Ca2+ steps. The origin of the Ca2+ steps and their role in the generation of long-lasting Ca2+ patterns were unclear. By simultaneous, noninvasive measuring of melanotrope plasma membrane electrical activity and the [Ca2+]i, we show that numbers, amplitude, and frequency of Ca2+ steps are variable among individual oscillations and are determined by the firing pattern and shape of the action currents. The general Na+ channel blocker tetrodotoxin had no effect on either action currents or the [Ca2+]i. Under Na+-free conditions, a depolarizing pulse of 20 mM K+ induced repetitive action currents and stepwise increases in the [Ca2+]i. The Ca2+ channel blocker CoCl2 eliminated action currents and stepwise increases in the [Ca2+]i in both the absence and presence of high K+. We furthermore demonstrate that the speed of Ca2+ removal from the cytoplasm depends on the [Ca2+]i, also between Ca2+ steps during the rising phase of an oscillation. It is concluded that Ca2+ channels, and not Na+ channels, are essential for the generation of specific step patterns and, furthermore, that the frequency and shape of Ca2+ action currents in combination with the Ca2+ removal rate determine the oscillatory pattern.

Action Potentials↗
PubMed1998

Cholinergic regulation of the pituitary: autoexcitatory control by acetylcholine of melanotrope cell activity in Xenopus laevis.

In the studies on Xenopus laevis presented here, the role of acetylcholine (ACh) in the regulation of pituitary activity was examined. The results are discussed in connection with other results on cholinergic regulation of pituitary functioning in vertebrates. It is demonstrated that dissociated melanotropes can synthesize ACh in vitro. In addition, immunocytochemical studies show the presence of the ACh-synthesizing enzyme choline acetyltransferase (ChAT) in the cytoplasm of melanotrope cells. ACh concentration-dependently raises the intracellular free calcium concentration as well as the release of proopiomelanocortin (POMC)-derived peptides. Selective muscarinic receptor antagonists showed that the actions of ACh are mediated through an M1-subtype muscarinic receptor. Immunofluorescence cytochemistry visualized muscarinic receptors on the surface of melanotrope cells. It is concluded that ACh stimulates the release of POMC-derived peptides from the Xenopus melanotrope cell in an autocrine, excitatory way. Finally, the possible mechanism through which ACh is released from melanotrope cells and the physiological significance of ACh in the pituitary gland are discussed.

Acetylcholinesterase↗
PubMed1998

Forebrain differentiation and axonogenesis in amphibians: I. Differentiation of the suprachiasmatic nucleus in relation to background adaptation behavior.

Functional forebrain development is the result of a complex series of early developmental processes which include cell division, cellular rearrangements, tissue-tissue interactions, cellular determinative and differentiation events, and axonogenesis. In these studies, Xenopus laevis embryos were examined for early forebrain neuronal determination, differentiation and axonogenesis with special emphasis on the hypothalamic area known to be involved in regulating pars intermedia function. Whole brain acetylcholine esterase (AChE) histochemistry was used to follow the early pattern of forebrain neuronal differentiation, and whole brain acetylated-tubulin immunocytochemistry was done to follow early forebrain axonogenesis. AChE histochemistry indicated that the source of the tract of the postoptic commissure (stpoc) was the first forebrain area to begin differentiation (stage 22). Whole brain immunocytochemistry for acetylated-tubulin indicated that the tpoc is also the first forebrain tract to develop (at stage 25/26). The main forebrain tracts have developed and become interconnected by stage 35/36. The forebrain undergoes a pronounced extension, with much cellular mixing and rearrangement during stages 37/38 to 43/44. This results in bending and contortions in the already developed tracts. Whole brain immunocytochemistry for tyrosine hydroxylase and extirpation of the stage 14 presumptive suprachiasmatic (SC) area indicated that the dopaminergic cells of the SC are determined by stage 14 and initially undergo differentiation between stages 37/38 and 40. Tadpoles with stage 14 presumptive SC extirpated lacked TH-positive tracts to the pars intermedia, lacked most midline TH-positive forebrain cells, and also failed to background adapt to white background. Thus, the SC tracts to the pars intermedia that inhibit melanotrope secretion probably form during the extension stages of 37/38 and contact the pars intermedia by stage 40 when animals are first capable of background adaptation.

Acetylcholine↗
PubMed1998

Dynamics of proopiomelanocortin and prohormone convertase 2 gene expression in Xenopus melanotrope cells during long-term background adaptation.

The toad Xenopus laevis is able to adapt its skin color to background light intensity. In this neuroendocrine reflex, the proopiomelanocortin (POMC)-derived peptide alpha-melanophore-stimulating hormone (alphaMSH) is a key regulatory factor. In animals adapting to a black background, release of alphaMSH from the pituitary pars intermedia causes dispersal of melanin in skin melanophores. To investigate the long-term in vivo dynamics of alphaMSH production during black background adaptation, the biosynthetic rate of POMC and the contents of POMC, alphaMSH and the POMC processing enzyme precursor convertase 2 (PC2) have been studied in the pars intermedia using pulse-labeling, Western blot and radioimmunoassay. In control animals, adapted to a white background, the rate of POMC biosynthesis and the POMC content were low, while high alphaMSH and PC2 contents were found. After 1 week of adaptation to a black background, the rate of POMC biosynthesis and the POMC protein content had increased 19- and 3.7-fold respectively. These parameters attained a maximum level (28- and 5. 8-fold higher than control) after 3 weeks and remained at these elevated levels for at least 12 weeks. After 1 week, the pars intermedia content of alphaMSH was only 30% of the control level, but after 6 and 12 weeks, the alphaMSH level had increased to the control level. The PC2 content decreased to 52% of control after 1 week and stabilized after 3 weeks at a level slightly lower than the control value. The results show that during long-term background adaptation a steady-state situation is reached, with a balance between the biosynthesis, enzymatic processing and release of alphaMSH. The in vivo dynamics of the processing enzyme PC2 suggest a parallel storage and release of alphaMSH and mature PC2 in the Xenopus pituitary pars intermedia.

Adaptation, Physiological↗
PubMed1997

Kinetics of calcium steps underlying calcium oscillations in melanotrope cells of Xenopus laevis.

Melanotrope cells of Xenopus laevis display intracellular calcium oscillations which are generated at the plasma membrane and travel as a wave through the cytoplasm into the nucleus. An oscillation involves discrete increases in intracellular Ca2+ ('steps'), followed by a relatively smooth return to the basal Ca2+ level. The aim of our investigation was to determine what role these steps play in shaping the Ca2+ signal in melanotrope cells, by conducting a high resolution spatio-temporal analysis of the kinetics of the Ca2+ steps. To this end Fura-red loaded cells were analysed by confocal laser scanning microscopy using the line scanning method to achieve 6 ms time resolution. Furthermore, the kinetics of the steps were analysed in 3 different intracellular areas, to see if there are spatial differences in Ca2+ signalling kinetics. The results showed that each calcium oscillation is built up by 3-4 steps that were generated very quickly and had approximately the same size. Following each Ca2+ step, there was a slow removal of calcium before the next step boosted the overall level of Ca2+. Since the Ca2+ steps were most pronounced directly beneath the plasma membrane, they appear to be generated in this region. The speed of the Ca2+ wave near the membrane exceeded 40 microns/s, indicating an active mechanism for wave propagation. In deeper regions of the cell, the wave speed was much slower (about 8 microns/s) and the size of each step was smaller, indicating that regulation occurs within a narrower range of [Ca2+]i. Inside the nucleus, however, the calcium wave accelerated again (23 microns/s). Treatment with TRH evoked a high amplitude Ca2+ transient and increased the number of Ca2+ steps to 5 or 6. Each step had approximately the same size as the steps of the pretreatment Ca2+ oscillations. Caffeine treatment, which increased the frequency of the oscillations, had no effect on the number or the size of the Ca2+ steps, but it reduced the time needed for each step to reach its maximum height. We suggest a possible 'building block' function for the Ca2+ steps, whereby a cell generates more steps to achieve a high oscillation amplitude or accelerates the speed of the steps to increase the frequency of oscillations. Both phenomena may play a crucial role in the encoding of information transduced from an extracellular input to the intracellular target.

Animals↗
PubMed1997

Nitric oxide synthase and background adaptation in Xenopus laevis.

Adaptation of the skin colour to the background light condition in the amphibian Xenopus laevis is achieved by migration of pigment granules in the skin melanophores, a process regulated by alpha-MSH secretion from melanotrope cells in the pituitary pars intermedia (PI). alpha-MSH secretion in turn, is regulated by various stimulatory and inhibitory messengers synthesized in brain nuclei, especially the hypothalamic suprachiasmatic and magnocellular nuclei and the locus coeruleus in the hindbrain. In the present study, the roles in background adaptation of nitric oxide (NO) and NO synthase (NOS) enzyme activity were evaluated. In situ, using both immunohistochemistry with anti-human brain NOS (bNOS) serum in paraffin-embedded material and using nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) histochemistry in cryo-sections, we showed NOS in neurons in the optic tectum and in the locus coeruleus. NADPH-d reactivity was also found in neurons in the lateral amygdala, the ventral hypothalamic nucleus and in fibers in the median eminence. Using a Western blot stained with an anti-human bNOS serum, we demonstrated a 150 kDa band in Xenopus hindbrain lysates, which is similar to the NOS protein present in the rat anterior pituitary, but which was not detectable in the lysates from both the neurointermediate and distal lobes in Xenopus. No differences in histochemical staining pattern or on Western blotting were observed between animals adapted to a black or a white background. Paraffin sections of the endocrine PI and pars distalis did not reveal bNOS-like immunoreactivity. NADPH-d reactivity was observed in the endothelia of this gland. However, using a new procedure of thin cryo-sections of pituitary neurointermediate lobes, we observed bNOS-immunoreactive fibers as well as cyclic 3',5' guanosine monophosphate (cGMP)-accumulating fibers in the PI. The PI may be regulated by NOergic neurons from higher brain centers. The possibility that NOergic neurons in the locus coeruleus are involved in the innervation of the PI needs further investigation. The latter neurons are probably not noradrenergic because double labeling studies show no co-localization of NADPH-d reactivity and tyrosine hydroxylase immunoreactivity in locus coeruleus neurons.

Adaptation, Physiological↗
PubMed1997

Physiologically induced Fos expression in the hypothalamo-hypophyseal system of Xenopus laevis.

The amphibian Xenopus laevis adjusts the color of its skin to the degree of background illumination. The neuroendocrine mechanism responsible for this adaptation behavior involves various brain centers that control the synthesis and release of alpha-melanophore-stimulating hormone (alpha-MSH) by the pituitary melanotrope cells. The aim of the present study was to investigate the possible use of Fos as a tool to determine the activity of known and novel components of this mechanism. For this purpose, a quantitative Fos-immunocytochemical method (ABC) was successfully introduced for Xenopus, and the degree of specificity of background illumination as a regulator of Fos expression was tested by comparing this stimulus with two other stimuli, viz. a strong stressor (saline immersion) and a mild stressor ('handling'). Without stimulation basal Fos-like immunoreactivity (Fos-LI) was found in the telencephalon, the lateral pallium, the anterior, central and lateral thalamic nuclei, the suprachiasmatic nucleus, the ventral hypothalamic nucleus and the torus semicircularis. Handling had no effect on this basal pattern of Fos-LI. Saline immersion induced Fos-LI only in the magnocellular preoptic nucleus, the corticotrope cells and, less strongly, in the melanotrope cells. Melanotropes, and no other cells, expressed Fos-LI very strongly when Xenopus was transferred from a white to a black background. This Fos-LI expression continued to increase up to 7 days of stimulation. When such toads were returned to a white background it took the same time before Fos-LI expression significantly dropped. It is concluded that the ABC-Fos immunocytochemistry can be successfully applied to assess the occurrence and degree of expression of Fos-LI in the Xenopus brain and pituitary gland. The prolonged expression of Fos-LI in the pars intermedia under black background stimulation and the presence of an AP-1 binding site on the Xenopus proopiomelanocortin gene suggest an important role for c-Fos and/or Fos-related antigens in the control of the biosynthesis and secretion of alpha-MSH by the Xenopus melanotrope cell.

Adaptation, Physiological↗
PubMed1997

Sauvagine and TRH differentially stimulate proopiomelanocortin biosynthesis in the Xenopus laevis intermediate pituitary.

In the amphibian Xenopus laevis, adaptation of the skin color to background light intensity is regulated by alpha-melanophore-stimulating hormone (alpha-MSH), a proopiomelanocortin (POMC)-derived peptide. In animals adapted to a white background, the level of POMC biosynthesis in the intermediate pituitary is much lower than in animals adapted to a black background. Release of alpha-MSH from neurointermediate lobes of white-adapted animals is stimulated in vitro by the regulatory peptides sauvagine and thyrotropin-releasing hormone (TRH), which are produced in the magnocellular nucleus of the hypothalamus. To study the role of sauvagine, cAMP, TRH and phorbol 12-myristate 13-acetate (PMA) in the regulation of POMC biosynthesis, the degree of incorporation of radioactive amino acids into the POMC protein was determined after treatment of the neurointermediate lobes with these secretagogues. When lobes of white-adapted animals are incubated in vitro, biosynthetic activity spontaneously increases because hypothalamic inhibitory control is removed by dissection. In addition to this control situation, the effects of secretagogues were tested on lobes with an inhibited level of biosynthesis, which is achieved by addition of neuropeptide Y (NPY) to the incubation medium. After 24 h of treatment, TRH stimulated POMC biosynthesis in NPY-inhibited lobes of white-adapted animals from 40.2 to 95.3% of control level. This stimulation could not be reduced by adding PMA, which indicates that protein kinase C is not involved in the stimulation of POMC biosynthesis by TRH. Sauvagine partially restored POMC biosynthesis from 27.2 to 62.5% of control level, whereas 8-Br-cAMP completely counteracted NPY inhibition from 27.8 to 97.5% of control level. After 3 days of treatment, stimulation by sauvagine and 8-Br-cAMP was maintained (sauvagine increased POMC biosynthesis in NPY-inhibited lobes from 7.4 to 36.2% of control level and 8-Br-cAMP stimulated from 6.5 to 82.5% of control level). TRH had no effect on POMC biosynthesis after 3 days of treatment, although its receptor was still functional as was shown in superfusion experiments where TRH stimulated alpha-MSH secretion. The observations indicate that the neuropeptides sauvagine and TRH differently control POMC biosynthesis in the Xenopus intermediate pituitary. This differential regulation is not only apparent with regard to time aspects (sauvagine has a sustained regulatory function, whereas TRH is only effective in the initial phase of POMC biosynthesis stimulation), but also an uncoupling of biosynthetic and release processes could be shown for TRH, which did not occur with sauvagine.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1997

Neuroendocrine gamma-aminobutyric acid (GABA): functional differences in GABAA versus GABAB receptor inhibition of the melanotrope cell of Xenopus laevis.

The melanotrope cell of Xenopus laevis is innervated by nerve terminals that contain, among other transmitter substances, the neurotransmitter gamma-aminobutyric acid (GABA). Postsynaptically the melanotrope cell possess both GABAA and GABAB receptors. Activation of either receptor type leads to an inhibition of alpha MSH release from the cell. The present study concerns the functional significance of the existence of two types of GABA receptors on the melanotrope regarding two questions: 1) do the different receptor types have different effects on the melanotrope? and 2) can the endogenous ligand GABA differentially activate these receptors? Concerning the first question, we have tested the hypothesis that the GABAA receptor (a chloride ion channel) and the GABAB receptor (a G protein-coupled receptor negatively linked to adenylyl cyclase) may have differential effects on the sensitivity of the cell to stimulation by cAMP-dependent mechanisms. We show that treatments with either isoguvacine (GABAA agonist) or baclofen (GABAB agonist) inhibit intracellular Ca2+ oscillations and peptide secretion from melanotrope cells. Treatments known to increase intracellular cAMP in the melanotrope (e.g. use of the peptide sauvagine or the cAMP analog 8-bromo-cAMP) completely overcame the inhibition induced by baclofen, but not that caused by isoguvacine. We conclude that the GABAA and GABAB receptors have different effects on the Xenopus melanotrope cell by differentially affecting the sensitivity the cell shows to stimulation by cAMP-dependent mechanisms. Concerning possible differential activation of the receptor types, we found that we could use a membrane potential probe (from the bis-oxonol family) to differentiate between GABAA and GABAB receptor activation. Using this probe we showed that low GABA concentrations (< 10(-7) M) give a response indicative of the GABAB receptor, whereas at high GABA concentrations (> 10(-7) M), the GABAA receptor response predominates. We, therefore, conclude that GABA can differentially activate the two types of GABA receptors on the Xenopus melanotrope cell.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1996

Identification of POMC processing products in single melanotrope cells by matrix-assisted laser desorption/ionization mass spectrometry.

The use of matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) in identifying proopiomelanocortin (POMC) processing products in melanotrope cells of the pituitary intermediate lobe of Xenopus laevis was explored. Mass spectra were obtained with such a high sensitivity of detection that the peptides could be identified in a single melanotrope cell. In addition to known POMC processing products of the Xenopus melanotrope cell, the presence of previously unidentified POMC-derived peptides was demonstrated. Together these POMC processing products accounted for the entire length of the POMC precursor. Furthermore, Xenopus possesses two genes for POMC and the sensitivity and accuracy of the MALDI-MS technique allowed identification of processing products of both the POMCA and POMCB gene. In addition, differences were obtained between the mass spectra of melanotrope cells from Xenopus laevis adapted to different conditions of background illumination. These results show that MALDI-MS is a valuable tool in the study of the expression of peptides in single (neuroendocrine) cells.

Amino Acids↗
PubMed1996

Spatial and temporal aspects of Ca2+ oscillations in Xenopus laevis melanotrope cells.

Spatio-temporal aspects of Ca2+ signaling in melanotrope cells of Xenopus laevis have been studied with confocal laser-scanning microscopy. In the whole-frame scanning mode, two major intracellular Ca2+ compartments, the cytoplasm and the nucleus, were visualized. The basal [Ca2+] in the nucleus appeared to be lower than that in the cytoplasm and Ca2+ oscillations seemed to arise synchronously in both compartments. The N-type channel blocker omega-conotoxin eliminated oscillations in both regions, indicating a strong coupling between the two compartments with respect to Ca2+ dynamics. Line-scanning mode, which gives higher time resolution, revealed that the rise phase of a Ca2+ oscillation is not a continuous process but consists of 3 or 4 discrete steps. Each step can be seen as a Ca(2+)-wave starting at the cell membrane and going through the cytoplasm at a speed of 33.3 +/- 4.3 microns/s. Before the Ca(2+)-wave enters the nucleus, a delay of 120.0 +/- 24.1 ms occurred. In the nucleus, the speed of a wave was 80.0 +/- 3.0 microns/s. Treatment with the Ca(2+)-ATPase inhibitor thapsigargin (1 MicroM) almost completely eliminated the apparent difference in the basal [Ca2+] in the cytoplasm and the nucleus, reduced the delay of a Ca(2+)-wave before entering the nucleus to 79.8 +/- 8.7 ms, and diminished the nuclear wave speed to 35.0 +/- 4.9 microns/s. These results indicate that a cytoplasmic thapsigargin-sensitive ATPase near the nuclear envelope is involved in buffering Ca2+ before the Ca2+ wave enters the nucleus. After sensitizing IP3 receptors by thimerosal (10 microM) the speed of the cytoplasmic Ca(2+)-wave was increased to 70.3 +/- 3.6 microns/s, suggesting that IP3 receptors may be involved in the propagation of the cytoplasmic Ca2+ wave. Our results indicate that in melanotropes the generation and propagation of Ca2+ oscillations is a complex event involving influx of Ca2+ through N-type Ca2+ channels, propagation of the cytoplasmic Ca2+ wave through mobilization of intracellular stores and a regulated Ca2+ entry into the nucleus. We propose that Ca(2+)-binding proteins may act as a Ca2+ store for propagation of the wave in the nucleus.

Animals↗
PubMed1996

Calcium oscillations in melanotrope cells of Xenopus laevis are differentially regulated by cAMP-dependent and cAMP-independent mechanisms.

Intracellular Ca2+ oscillations play an important role in the induction of alpha-MSH release from pituitary melanotrope cells of Xenopus laevis. Oscillatory, secretory and adenylyl cyclase activities are all inhibited by dopamine, neuropeptide Y (NPY) and baclofen (a GABAB receptor agonist) and stimulated by sauvagine. In this study, we test the hypothesis that these neural messengers regulate the Ca2+ oscillations via a cAMP/protein kinase A (PKA)-dependent mechanism. To this end, video-imaging microscopy was applied to single Xenopus melanotropes loaded with the Ca2+ indicator Fura-2. The cAMP-dependent PKA inhibitor H89 blocked Ca2+ oscillations as well as the stimulatory actions of 8-Br-cAMP and sauvagine. Treatment of cells inhibited by baclofen with either 8-Br-cAMP or sauvagine led to a reappearance of Ca2+ oscillations. A similar result was found for cells inhibited by NPY. Neither 8-Br-cAMP nor sauvagine induced Ca2+ oscillations in cells inhibited by dopamine. Depolarizing dopamine-inhibited cells with high potassium also failed to induce oscillations, but combining 8-Br-cAMP with membrane depolarization induced oscillations. It is concluded that sauvagine, baclofen and NPY work primarily through a cAMP/PKA-pathway while dopamine inhibits Ca2+ oscillations in a dual fashion, namely via both a cAMP-dependent and a cAMP-independent mechanism, the latter probably involving membrane hyperpolarization.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1996

Acetylcholine autoexcites the release of proopiomelanocortin-derived peptides from melanotrope cells of Xenopus laevis via an M1 muscarinic receptor.

The release of POMC-derived peptides from the melanotrope cells in the pituitary pars intermedia of Xenopus laevis is regulated by multiple inhibitory and stimulatory neurochemical messengers. In this study we examined the role of acetylcholine (ACh) in regulating melanotrope cell activity. The origin of ACh was studied in in vitro biosynthesis experiments, where it was demonstrated that dissociated melanotropes can synthesize ACh. In addition, immunocytochemical studies showed the presence of the ACh-synthesizing enzyme choline acetyltransferase in the cytoplasm of melanotrope cells. Oscillations in the concentration of intracellular free calcium ions are thought to be the driving force for the secretion of POMC-derived peptides from Xenopus melanotrope cells. A technique was developed to study simultaneously calcium oscillations and peptide secretion from the same population of single melanotrope cells in vitro. ACh concentration-dependently raises the intracellular free calcium concentration by increasing both the frequency and amplitude of the calcium oscillations as well as the release of POMC-derived peptides. The actions of ACh on the melanotrope cell were mimicked by muscarine, but not by nicotine, indicating the exclusive presence of a muscarinic cholinergic receptor. Furthermore, selective muscarinic receptor antagonists showed that the actions of ACh are mediated through an M1-subtype muscarinic receptor. Immunofluorescence cytochemistry in combination with confocal laser scanning microscopy visualized muscarinic receptors on the surface of melanotrope cells. It is concluded that ACh stimulates the release of POMC-derived peptides from the Xenopus melanotrope cell in an autocrine way, acting via a muscarinic M1 receptor on the intracellular free calcium concentration.

Acetylcholine↗