Biomedical subjects

B G Jenks

Publications and source records attributed to B G Jenks.

At least 73 records · Page 4Linked to original sources

PubMed1993

Effect of dopamine on adenylate cyclase activity, polyphosphoinositide metabolism and cytosolic calcium concentrations in frog pituitary melanotrophs.

It has previously been shown that dopamine plays a pivotal role in the regulation of alpha-melanocyte-stimulating hormone (alpha-MSH) secretion from the intermediate lobe of the pituitary. In the present study, we have investigated the various intracellular mechanisms that are associated with the action of dopamine on frog pituitary melanotrophs. Dopamine reduced forskolin-stimulated cyclic adenosine monophosphate (cAMP) production and the inhibitory effect of dopamine was blocked by the dopaminergic D2 receptor antagonist sulpiride. The D2 receptor agonist apomorphine inhibited incorporation of [3H]inositol into membrane phospholipids. Dopamine also inhibited the formation of inositol trisphosphate and provoked accumulation of phosphatidylinositol bisphosphate. The inhibitory effect of dopamine on inositol trisphosphate production was mimicked by D2 receptor agonists and blocked by sulpiride. Using a double-wavelength microfluorimetric approach, we found that dopamine caused a rapid and transient decrease in K(+)-evoked stimulation of intracellular calcium concentration. The time-courses of the responses of the various intracellular messengers indicate that blockage of voltage-dependent calcium channels is the primary event associated with activation of dopamine D2 receptors, while inhibition of polyphosphoinositide breakdown, related to blockage of voltage-dependent calcium channels, and reduction of cAMP production are secondary events which may contribute to the sustained inhibitory effect of dopamine on alpha-MSH release.

Adenylyl Cyclases↗

PubMed1992

Immunoblotting technique to study release of melanophore-stimulating hormone from individual melanotrope cells of the intermediate lobe of Xenopus laevis.

The melanotrope cells in the pars intermedia in the pituitary of Xenopus laevis synthesize and release the melanophore-stimulating hormone (alpha MSH), a small peptide that causes skin darkening during the process of background adaptation. Evidence has been found for a heterogeneity in biosynthetic activity of the melanotrope cells. In the present study two questions were addressed: (1) does the melanotrope cell population also show heterogeneous alpha MSH-release, and (2) can this heterogeneity be changed by extracellular messengers? Since dopamine is known to inhibit alpha MSH-release, this messenger is used to study the regulation of the heterogeneity. To quantify alpha MSH-release from individual cells, a cell blotting procedure has been developed for the binding and relative quantification of the small alpha MSH peptide. The immunoblotting procedure involves binding of the cells to a carrier slide and binding of released alpha MSH to a nitrocellulose filter. After immunostaining, the amount of alpha MSH per cell was quantitated by image analysis. Untreated melanotrope cells reveal a distinct variability in alpha MSH-release, some cells showing low secretory activity, whereas others are strongly secreting, indicating heterogeneity of alpha MSH-release. Dopamine treatment strongly inhibits alpha MSH-release from individual cells, resulting in a clearly less pronounced melanotrope cell heterogeneity. The effect of dopamine appears to be dose-dependent as a low dopamine concentration has only a moderate effect on the alpha MSH-release. It is proposed that dopamine is a physiological regulator of the degree of melanotrope cell heterogeneity in alpha MSH-release.

Animals↗

PubMed1992

Analysis of autofeedback mechanisms in the secretion of pro-opiomelanocortin-derived peptides by melanotrope cells of Xenopus laevis.

The secretion of most pituitary hormones is under the control of feedback mechanisms. The feedback control of alpha-melanophore-stimulating hormone (alpha-MSH) from melanotrope cells is controversial. The possible existence of an autofeedback exerted by alpha-MSH or other POMC-derived peptides on melanotrope cells of the amphibian Xenopus laevis has been investigated. alpha-MSH or its potent agonist 4-norleucine,7-D-phenylalanine-alpha-MSH has no effect on the release of radiolabeled POMC-derived peptides or immunoreactive beta-endorphin from superfused neurointermediate pituitary lobes. Melanin concentrating hormone, previously reported to have an alpha-MSH-like effect on melanophores, did not affect alpha-MSH secretion. Neurointermediate lobe superfusate, which contains a mixture of POMC-derived peptides, failed to affect the secretory activity of melanotropes. It is concluded that in X. laevis the secretory activity of melanotropes is not under the control of short-term autofeedback mechanisms involving alpha-MSH or other POMC-derived peptides.

Animals↗

PubMed1992

Dynamics of cyclic-AMP efflux in relation to alpha-MSH secretion from melanotrope cells of Xenopus laevis.

An important factor in regulating secretion from endocrine cells is the cytoplasmic concentration of cyclic-AMP. Many regulatory substances are known to either stimulate or inhibit the production of this second messenger through activation of their receptors. In the present study, we have monitored changes in cyclic-AMP efflux from melanotrope cells of Xenopus laevis in response to established neurochemical regulators of alpha-MSH secretion. In vitro superfusion of neurointermediate lobes allows for a dynamic recording of cyclic-AMP production in relation to hormone secretion. Unlike alpha-MSH secretion, the efflux of cyclic-AMP was not dependent on the concentration of extracellular calcium, indicating that hormone release and cyclic-AMP efflux are mediated by different mechanisms. The phosphodiesterase inhibitor IBMX and the adenylate cyclase activator forskolin stimulated cyclic-AMP efflux, but had no stimulatory effect on alpha-MSH release. This indicates that an increase in cyclic-AMP production in melanotrope cells is not necessarily accompanied by an increase in the rate of alpha-MSH release. Corticotropin-releasing factor stimulated cyclic-AMP efflux with dynamics similar to that induced by the amphibian peptide sauvagine. Dopamine and the GABAB receptor agonist baclofen both inhibited cyclic-AMP efflux and alpha-MSH release, with similar dynamics of inhibition and similar dose-response relationships. It is proposed that an inhibition of cyclic-AMP efflux is coupled to an inhibition of alpha-MSH secretion.

1-Methyl-3-isobutylxanthine↗

PubMed1992

Transcriptional and posttranscriptional regulation of the proopiomelanocortin gene in the pars intermedia of the pituitary gland of Xenopus laevis.

In the melanotrope cells of the intermediate pituitary gland of the amphibian Xenopus laevis, expression of the POMC gene is under physiological control, i.e. it is 20- to 30-fold higher in animals adapted to a black background compared to animals adapted to a white background. To investigate whether changes in POMC messenger RNA stability contribute to this difference in expression, a steady state kinetic model for mRNA degradation was used to determine the half-life of POMC mRNA during the induction and deinduction of POMC gene expression in melanotrope cells. During induction of the POMC gene the half-life of POMC mRNA was 3- to 4-fold longer than during deinduction. This difference in mRNA stability is, however, not sufficient to account for the 20- to 30-fold difference in the steady state levels of POMC mRNA between the two physiological conditions. Results from experiments with the protein synthesis inhibitor cycloheximide and the mRNA synthesis inhibitor 5,6-dichloro-1-beta-D-ribofuranosylbenzidimidazole suggest 1) that during induction of POMC gene expression no de novo protein synthesis is required and, 2) that deinduction of POMC gene expression requires transcriptional activation of an mRNA-degradation system. Pulse-labeling experiments with [3H]uridine showed that in neurointermediate lobes of white-adapted animals there is an 8-fold higher amount of newly synthesized POMC mRNA than in lobes of black animals. This suggests a fast in vitro induction of POMC gene transcription. The dopamine D2-receptor agonist apomorphine decreased POMC mRNA biosynthesis about 5-fold, which confirms that regulation of POMC gene expression includes a transcriptional component. In conclusion, our results demonstrate that during the physiological process of background adaptation the regulation of POMC gene expression in Xenopus melanotrope cells is exerted by alterations in POMC gene transcription as well as in POMC mRNA stability.

Animals↗

PubMed1991

A genetic-correlational study of hippocampal structural variation and variation in exploratory activities of mice.

Our previous work provided evidence that hippocampal opioid peptides form an important neurochemical substrate underlying the gene-dependent exploratory behavior of mice. A prominent hippocampal opioid is dynorphin B, which resides in the mossy fibers exclusively. In order to seek support for causal relationships between dynorphinergic hippocampal mechanisms and exploration, a quantitative-genetic method was chosen. For this purpose, mice from the inbred strains C57BL/6, DBA/2, BLN, and CPB-K were used. Their hippocampal mossy fiber projections were visualized by means of immunohistochemistry, using a highly specific anti-dynorphin B antiserum. The additive-genetic correlations that were estimated suggest pleiotropic gene effects on locomotion, rearing-up, wall-leaning, and several intra- and infrapyramidal mossy fiber (iipMF) variables. Long iipMF, in particular, were found to be associated with high exploratory activity.

Animals↗

PubMed1991

A genetic-correlational study of hippocampal neurochemical variation and variation in exploratory activities of mice.

Previously, we have demonstrated that hippocampal mossy fibers, containing the opioid peptide dynorphin B, are functionally connected with the gene-dependent exploratory behavior of mice. In order to seek further evidence of causal relationships between dynorphin B action and exploration, a quantitative-genetic method was chosen. For this purpose, mice from the inbred strains C57BL/6, DBA/2, BLN, and CPB-K were used. By means of radioimmunoassay, the hippocampal level of dynorphin B was monitored in mice that had been exposed to environmental novelty, as compared to naive animals. Clear evidence was obtained that novelty induces the release of hippocampal dynorphin B. Furthermore, low tissue content was found to be causally connected with high exploratory scores.

Animals↗

PubMed1991

Coordinated expression of 7B2 and alpha MSH in the melanotrope cells of Xenopus laevis. An immunocytochemical and in situ hybridization study.

7B2 is a highly conserved protein present in many secretory cells. Using in situ hybridization techniques and immunocytochemistry, parameters concerning the biosynthesis and storage of the 7B2 protein were studied in the pituitary gland and median eminence of the clawed toad Xenopus laevis, in relation to the physiological process of background adaptation. 7B2-like immunoreactivity was present in the median eminence, in the neural and anterior pituitary lobes and, particularly, in the melanotrope cells of the intermediate pituitary lobe. In these cells, it coexisted with immunoreactivity to proopiomelanocortin (POMC)-derived alpha-melanocyte stimulating hormone (alpha MSH). The melanotropes of black-adapted animals had abundant 7B2-mRNA and POMC-mRNA; melanotropes of white-adapted toads had only low levels of these mRNAs. The presence of 7B2 in nerve terminals and endocrine cells supports the idea that the protein has a general function in the cellular secretory process. In X. laevis, 7B2 appears to be particularly associated with POMC and alpha MSH and, therefore, may play a role in the regulation of background adaptation.

Animals↗

PubMed1991

The CRF-related peptide sauvagine stimulates and the GABAB receptor agonist baclofen inhibits cyclic-AMP production in melanotrope cells of Xenopus laevis.

Release of alpha-MSH from the pars intermedia melanotrope cells of Xenopus laevis is regulated by various classical neurotransmitters and neuropeptides. We have examined the effect of two of these regulatory substances, the neurotransmitter GABA and the CRF-related peptide sauvagine, on the adenylate cyclase system of the melanotrope cells. Sauvagine treatment, which stimulates alpha-MSH release, lead to an elevation in the level of cyclic-AMP, an effect which was potentiated by cholera toxin. Treatment with baclofen, a GABAB receptor agonist, gave a pertussis toxin-sensitive decrease in the cyclic-AMP level and an inhibition of alpha-MSH release. We conclude that sauvagine stimulates alpha-MSH secretion through activation of adenylate cyclase and that GABAB receptor activation inhibits secretion through inhibition of cyclic-AMP production. Baclofen treatment sensitized melanotrope cells to the stimulatory action of 8-bromo-cyclic-AMP on the secretion of alpha-MSH. This observation supports the conclusion that GABAB receptor activation inhibits cyclic-AMP production.

Adenylyl Cyclases↗

PubMed1991

Indirect action of elevated potassium and neuropeptide Y on alpha MSH secretion from the pars intermedia of Xenopus laevis: a biochemical and morphological study.

A number of neurochemical messengers have been shown to act directly on the melanotrope cells of the pars intermedia of Xenopus laevis to regulate alpha MSH secretion. In the present study the possibility that the melanotropes are also indirectly controlled has been examined. For this purpose, the characteristics of alpha MSH release from superfused intact lobes, cultured lobe and isolated melanotropes were compared after treatment with elevated potassium. Isolated melanotropes responded with an increased secretion of alpha MSH, whereas intact lobes showed a profound inhibitory response, probably caused by potassium-induced release of inhibitory factors from nerve terminals. Cultured lobes displayed a biphasic response characterized by an initial activation followed by a strong inhibition; the stimulatory phase likely reflects a direct action of potassium on the melanotropes, before being overridden by an inhibitory mechanism. The inhibitory phase must originate from the action of nonneuroendocrine cells because the cultured lobes lack functionally active nerve terminals, as verified by immunocytochemistry and electron microscopy. The most likely candidates for this action are folliculo-stellate cells which are in intimate contact with the melanotropes and are innervated by neuropeptide Y-containing nerve terminals. Like elevated potassium, neuropeptide Y inhibited alpha MSH secretion from fresh and cultured lobes but not from isolated melanotropes. This indicates that NPY acts indirectly, in a nonpresynaptic way, to inhibit alpha MSH secretion.

Animals↗

PubMed1991

[125I]Bolton-Hunter neuropeptide-Y-binding sites on folliculo-stellate cells of the pars intermedia of Xenopus laevis: a combined autoradiographic and immunocytochemical study.

It has previously been established that neuropeptide-Y (NPY) is a potent inhibitor of alpha MSH release from the pars intermedia of the amphibian Xenopus laevis. The location of binding sites for NPY in the pars intermedia of the pituitary has now been studied with light microscopic autoradiography, using a dispersed cell labeling method with the specific NPY receptor ligand [125I]Bolton-Hunter NPY. The majority of radioactive labeling was associated with folliculo-stellate cells; the percentage of labeling as well as the mean number of grains were approximately 5 times higher for folliculo-stellate cells than for melanotropes. An excess of nonlabeled NPY drastically reduced radiolabeling of folliculo-stellate cells, but had no effect on the degree of labeling of melanotropes. These results show that folliculo-stellate cells of X. laevis possess specific binding sites for NPY and indicate that NPY exerts its inhibitory action on the release of alpha MSH in an indirect fashion, by acting on the folliculo-stellate cells.

Animals↗

PubMed1991

Regulation of differential release of alpha-melanocyte-stimulating hormone forms from the pituitary of a teleost fish, Oreochromis mossambicus.

Using high-performance liquid chromatography (HPLC) in combination with radioimmunoassay, three forms of alpha-MSH (des-acetyl, mono-acetyl and di-acetyl alpha-MSH) were separated and identified in tilapia neurointermediate lobes and plasma, and in medium from lobes superfused in vitro. The presence of acetylated forms in lobe extracts indicated that the peptides are acetylated intracellularly. Di-acetyl alpha-MSH was, especially in comparison with monoacetyl alpha-MSH, relatively more abundant in lobe extracts than in plasma. This suggests that the three forms of alpha-MSH are not released according to their relative intracellular abundances. The possibility of regulation of this differential release by dopamine and TRH was investigated, using a microsuperfusion system. Dopamine was a potent inhibitor of alpha-MSH release, but did not modulate the relative abundance of the different forms of alpha-MSH released from the MSH cells. TRH was a potent stimulator of alpha-MSH release. It enhanced in vitro the release of di-acetyl alpha-MSH more than the release of mono-acetyl alpha-MSH. Thus tilapia may be able to modulate not only the quantitative but also the qualitative signal from the MSH cells. This might enhance the flexibility of the animals to respond to environmental challenges.

Animals↗

PubMed1990

The neuroendocrine polypeptide 7B2 is a precursor protein.

The neuroendocrine protein 7B2 is highly conserved and widely present in neurons and endocrine cells. It is coexpressed with the prohormone proopiomelanocortin (POMC) in the intermediate lobe of the pituitary gland of Xenopus laevis. To study the biosynthesis of 7B2 in this amphibian, an anti-7B2 monoclonal antibody was used in immunoprecipitation analysis of newly synthesized radiolabeled proteins, produced by pulse and pulse-chase-incubated neurointermediate lobes. Following a 15-min pulse incubation, a single immunoprecipitable protein of 25 kDa was synthesized. During subsequent chase incubation, this newly synthesized 7B2 protein was processed to an 18-kDa immunoprecipitable form. Analysis of the chase incubation medium revealed that only the 18-kDa processed product of 7B2, and not 7B2 itself, had been secreted. This secretion is a regulated process because it was blocked completely by the dopamine receptor agonist apomorphine. A study of protein biosynthesis in lobes treated with tunicamycin to prevent N-linked glycosylation showed that in contrast to POMC and an 18-kDa derivative of POMC, neither 7B2 nor its 18-kDa derivative was glycosylated. Chemical and enzymatic peptide mapping showed that processing of 7B2 occurs in the carboxyl-terminal region. The function of the 7B2 protein is unknown; the present results show that 7B2 itself is a precursor molecule and can only have an intracellular function whereas an extracellular function can only be attributed to 7B2-derived peptides.

Amino Acid Sequence↗

PubMed1990

Electrophysiological evidence for the existence of GABAA receptors in cultured frog melanotrophs.

The neurotransmitter GABA exerts a biphasic effect on alpha-melanocyte-stimulating hormone (alpha-MSH) secretion from pars intermedia cells: GABA induces a rapid and transient stimulation followed by a sustained inhibition of alpha-MSH release. In the present study, we have investigated the effect of GABA on the electrophysiological properties of frog melanotrophs in primary culture using the patch-clamp technique in the whole cell configuration. In all cells tested, GABA stimulated an inward current and induced depolarization. A transient period of intense firing was consistently observed at the onset of GABA administration. During the depolarization phase, the membrane potential reached a plateau corresponding to the Cl- equilibrium potential. When repeated hyperpolarizing pulses were applied, an increase of membrane conductance was observed throughout the response evoked by GABA. The effect of GABA was abolished by the chloride channel blocker picrotoxin, and by antagonists of GABAA receptors (bicuculline and SR 95531). The depolarizing action of GABA was mimicked by muscimol, an agonist of GABAA receptors. Taken together, our results indicate that the rapid and transient stimulation of alpha-MSH release induced by GABA can be accounted for by activation of a chloride conductance which causes membrane depolarization. These data support the notion that the transient stimulation of alpha-MSH secretion induced by GABA can be accounted for by membrane depolarization which provokes activation of voltage-operated calcium channels. Since no evidence was found for GABA-induced hyperpolarization, the intracellular mechanisms leading to the strong inhibitory effect of GABA on alpha-MSH secretion remain to be elucidated.

Animals↗

PubMed1990

Morphology of the pars intermedia and the melanophore-stimulating cells in Xenopus laevis in relation to background adaptation.

The melanophore-stimulating hormone (MSH) cells of the amphibian pars intermedia secrete the peptide alpha-melanophore-stimulating hormone (alpha-MSH), which induces pigment dispersion in dermal melanophores. The purpose of the present study was to determine (1) whether prolonged activation of the secretory activity of the pars intermedia is associated with hypertrophy, hyperplasia, or both and (2) whether the MSH cells function as a homogeneous or heterogeneous population in meeting the physiological demand for MSH. The demand for MSH was manipulated by adapting animals for at least 3 weeks to white, two shades of grey, or black backgrounds. Morphometric analysis showed that the intermediate lobe volume was positively correlated with the degree of pigment dispersion in the melanophores. The number of MSH cells per lobe was not affected by the degree of pigment dispersion. Therefore, we conclude that enlargement of the tissue associated with MSH cell activation involves hypertrophy rather than hyperplasia. Ultrastructural examination indicated that the majority of MSH cells in black-adapted animals are biosynthetically active, whereas the cells of white-adapted animals are relatively inactive and show granule storage. The pars intermedia of grey-adapted toads contained both active and inactive cells, indicating that MSH cells respond as a heterogeneous cell population in meeting the endocrine demand imposed by background.

Adaptation, Biological↗

PubMed1990

Neuropeptide Y inhibits thyrotropin-releasing hormone-induced stimulation of melanotropin release from the intermediate lobe of the frog pituitary.

Previous studies have shown that the release of melanotropin from frog neurointermediate lobes is under the control of two neuropeptides: thyrotropin-releasing hormone (TRH) stimulates, while neuropeptide Y (NPY) inhibits alpha-melanocyte-stimulating hormone (alpha-MSH) secretion from intact neurointermediate lobes in vitro. The aim of the present study was to investigate possible interactions between the two regulatory peptides at the pituitary level. Whole neurointermediate lobes or acutely dispersed pars intermedia cells from Rana ridibunda were perifused in vitro for 2 to 7.5 hr and the concentrations of alpha-MSH released into the effluent perifusate were monitored by radioimmunoassay. Administration of TRH (10(-7) M) or NPY (10(-7) M) to dispersed cells induced, respectively, marked stimulation or inhibition of alpha-MSH release. The effects of the two neuropeptides were similar to those observed using intact neurointermediate lobes, suggesting that TRH and NPY act directly on melanotropic cells. Perifused whole neurointermediate lobes were exposed to NPY (10(-8) to 3 x 10(-7) M) for 120 min and a single dose of TRH (10(-8) M) was administered during the prolonged infusion of NPY. Using this procedure, we observed a dose-dependent inhibition of TRH-evoked alpha-MSH release. These data support the concept that TRH and NPY act through a common intracellular pathway to regulate alpha-MSH release.

Animals↗

PubMed1990

GABA and neuropeptide Y co-exist in axons innervating the neurointermediate lobe of the pituitary of Xenopus laevis--an immunoelectron microscopic study.

The neural innervation of the neurointermediate lobe of the pituitary of the amphibian Xenopus laevis has been studied at the light and electron microscopic level. In the pars intermedia melanotropes and stellate cells are abutted by varicosities originating from GABA- and neuropeptide Y-producing neurons. The varicosities contain two types of vesicle: electron-lucent vesicles (mean diameter 50 nm) which are immunopositive for GABA and larger (80 nm) electron-dense vesicles which are immunopositive for neuropeptide Y. Double immunogold labeling established that GABA and neuropeptide Y co-exist within the varicosities. In the pars nervosa similar varicosities, though low in number, occur. They are associated with neurosecretory nerve terminals, pituicytes and blood vessels. The possible significance of GABA and neuropeptide Y for the neural regulation of melanophore stimulating hormone-release from the pars intermedia is discussed.

Animals↗

PubMed1990

A slow and a fast secretory compartment of POMC-derived peptides in the neurointermediate lobe of the amphibian Xenopus laevis.

1. Peptide release from the neurointermediate lobe of Xenopus laevis has been studied using dual pulse-chase incubation, superfusion and HPLC techniques. 2. Lobes release pulse-labelled material in two phases, the first phase lasting about 6 hr, the second persisting up to 14 hr. 3. In both phases similar, POMC-derived peptides are released. Their release can be inhibited by dopamine. 4. When release during the first phase is inhibited, newly synthesized peptides are shunted into the second release pathway. 5. It is concluded that the neurointermediate lobe contains two release compartments. The possible locations of these compartments within melanotrope cells have been discussed.

Adrenocorticotropic Hormone↗