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B M Chronwall

Publications and source records attributed to B M Chronwall.

At least 37 records · Page 2Linked to original sources
PubMed1995

Glial-like cells of the rat pituitary intermediate lobe change morphology and shift from vimentin to GFAP expression during development.

This study demonstrated morphological changes in glial-like cells of the rat pituitary intermediate lobe during early postnatal development, and a subsequent shift in protein expression from vimentin to GFAP. Vimentin immunoreactivity was detected in the lobe at embryo day 14 and was localized in radially-oriented, bipolar cells whose processes spanned the thickness of the intermediate lobe. At electron microscopical resolution, processes contained intermediate filaments, cell nuclei were indented while secretory vesicles characteristic of the endocrine cells were not found. Vimentin immunoreactive intensity began to decrease at postnatal day 5. By postnatal day 7, vimentin-positive, stellate cells were observed, with few radial processes found by day 10. The intensity of vimentin immunoreactivity decreased through day 25. Within the lobe parenchyma, vimentin was localized in glial-like cells since double-label immunohistochemistry revealed no colocalization of beta-endorphin and vimentin, or fibronectin and vimentin. Dopamine-containing axons were in close apposition to vimentin-positive processes. GFAP immunoreactivity first appeared on postnatal day 20 and, by day 25, stellate cell bodies with three to six extended processes were evident. Cells were primarily distributed in the caudal third of the lobe. The characteristic adult pattern of cell clusters in latero-dorsal and ventral portions of the lobe was fully established by postnatal day 55. The transition from vimentin to GFAP expression and concurrent morphological changes resemble those described for radial glial during cerebral cortical development.

Animals↗
PubMed1995

Transient expression of S-100 by melanotropes of the rat pituitary intermediate lobe during development.

S-100, an acidic calcium-binding protein, is co-localized with vimentin in glial-like cells in the adult rat pituitary intermediate lobe. S-100 and melanotrope markers were not co-localized in the adult. During development, S-100 and vimentin were not co-localized but appeared in cells with different morphological characteristics. S-100 was co-localized with POMC mRNA and beta-endorphin during prenatal time and the first three postnatal weeks. This was demonstrated by double-label immunohistochemistry, using combinations of antisera against S-100, vimentin and beta-endorphin, and in situ hybridization histochemistry for POMC mRNA combined with immunohistochemistry for S-100. In the second and third weeks of postnatal development, S-100 was observed in fewer melanotropes and more frequently in stellate cells, which also expressed vimentin. Thus, S-100 appeared to be transiently expressed in melanotropes during prenatal and early postnatal development. S-100 serves as a neurotrophic and glial maturation factor in the CNS. Since S-100 expression in melanotropes coincides with the onset of dopaminergic innervation and morphological changes in glial-like cells of the lobe, it could have similar functions in the rat pituitary intermediate lobe.

Animals↗
PubMed1995

Dopamine D2 receptors regulate in vitro melanotrope L-type Ca2+ channel activity via c-fos.

Dopamine D2 receptor stimulation of cultured primary melanotropes was found to depress L-type calcium channel activity, whereas D2 receptor antagonist application increased it. When tested on culture days 10, 16, and 20, control cells displayed increasing rises of intracellular Ca2+ in response to K+ depolarization, indicating an increase in channel activity in the absence of dopaminergic regulation. When treated with 1 microM bromocriptine from culture day 1, cells showed minimal increase in channel activity. When bromocriptine was added on day 16, intracellular Ca2+ response to high K+ declined by day 20; removal of the agonist on day 16 resulted in the reappearance of increased responsiveness. Thus, in vitro inhibitions could be initiated or reversed with application or withdrawal of dopamine D2 receptor agonist. Cultured melanotropes were treated with antisense oligodeoxynucleotides directed against the start sequences of the D2 receptor and c-fos messenger RNA. D2 receptor antisense nucleotide prevented the depressive effect on channel activity induced by D2 agonist treatment. c-fos antisense oligodeoxynucleotide blocked the rise in channel activity. The dopamine D2 receptor antagonist haloperidol, which increased channel activity, could not reverse the c-fos antisense deoxynucleotide block. These results strongly support the idea that the chronic suppression of secretion-related activities by dopaminergic stimulation seen in the intermediate lobe in vivo is effected by chronic suppression of c-fos by D2 receptors.

Animals↗
PubMed1994

Regulation of heterogeneity in D2 dopamine receptor gene expression among individual melanotropes in the rat pituitary intermediate lobe.

The regulation of D2 dopamine receptor binding, immunoreactivity, and mRNA was studied in melanotropes of the intermediate lobe of the rat pituitary. D2 dopamine receptor-binding experiments showed that chronic haloperidol treatment (2 mg/kg for 14 days) significantly increased the Bmax but not Kd of [3H]spiperone binding, whereas bromocriptine treatment (2 mg/kg for 14 days) produced no change in either parameter. Immunohistochemical experiments revealed that all melanotropes contained D2 receptor immunoreactivity but the D2long receptor isoform was intensely expressed by comparatively few melanotropes scattered among cells that contained substantially lower staining intensities. Chronic haloperidol treatment increased the overall intensity of total D2 receptor staining and concurrently increased both the number and staining intensity of melanotropes immunoreactive for the D2long isoform. Bromocriptine treatment produced no change in overall staining intensity; however, the number of melanotropes staining for the D2long isoform increased significantly. Nonradioactive in situ hybridization histochemistry further revealed that individual melanotropes contained differing amounts of both total D2 receptor and D2long mRNAs; the heterogeneity in D2long receptor expression was particularly striking. Following haloperidol treatment, D2 receptor mRNA levels increased in all melanotropes. Following bromocriptine treatment most melanotropes contained very low levels although many melanotropes retained substantial amounts of D2 receptor mRNA. The parallel increase in D2 receptor-binding densities, immunoreactivity, and mRNA levels following chronic antagonist treatment emulates the classic paradigm by which a cell increases its receptor population. Chronic agonist treatment did not follow the inverse paradigm and revealed heterogeneous regulation of a discrete subpopulation of melanotropes.

Amino Acid Sequence↗
PubMed1994

POMC mRNA levels in individual melanotropes and GFAP in glial-like cells in rat pituitary.

In this study we demonstrated that individual melanotropes expressed differing levels of POMC mRNA. Interspersed among the melanotropes was a small population of process-bearing cells expressing GFAP. We compared cellular resolution and feasibility for quantitation of hybrids formed by oligonucleotide probes labeled with two different markers using in situ hybridization histochemistry. In hybridizations using [35S]-labeled probes the signal could not easily be assigned to individual melanotropes, although digoxigenin-labeled probes provided good cellular resolution. Heterogeneous staining intensities of individual melanotropes for POMC mRNA were measured, and relative quantitation of changes in POMC mRNA levels following interactions with the D2 dopamine receptor was performed. We combined in situ hybridization histochemistry with immunohistochemistry to demonstrate that cells within the lobe parenchyma not expressing POMC mRNA were immunoreactive for GFAP.

Animals↗
PubMed1994

Localization of neuropeptide Y immunoreactivity and [125I]peptide YY binding sites in the human pituitary.

High levels of neuropeptide Y (NPY) are found in the hypothalamus, median eminence, pituitary portal blood, and the pituitary in a number of species. Neuropeptide Y may influence the synthesis and secretion of a variety of hormones by interacting with specific receptors in the hypothalamus and/or the pituitary. To further define the function of NPY in the pituitary, we have examined the distribution of NPY immunoreactivity and NPY receptors in sections of human pituitary using immunohistochemical and autoradiographic techniques. Neuropeptide Y-immunoreactive varicose axons were seen throughout the neural lobe. A moderate number of NPY-immunoreactive cells were found in the anterior lobe. A very high level of [125I]PYY binding was seen in the neural lobe with low levels in the anterior lobe. The binding in the neural lobe was inhibited by NPY(13-36) at a Ki of 5.3 nM and [Leu31-Pro34]NPY at a Ki of 390 nM, indicating the receptor was the Y2 subtype. Therefore, neuronally released NPY may modulate human neural lobe function through a Y2 receptor.

Adolescent↗
PubMed1994

Development of neuropeptide Y and tyrosine hydroxylase immunoreactive innervation in postnatal rat heart.

Neuropeptide Y (NPY), immunoreactive (IR), and tyrosine hydroxylase (TH)-IR nerve fibers were scarce at birth in rat heart, but increased rapidly during the first 2 postnatal weeks, reaching approximately adult levels by the third week. The sequence of development was: interatrial septum and atrial wall, free ventricular wall starting from the epicardium, and finally the atrial appendages and interventricular septum. In ventricles and atrial appendages both fiber types developed similarly. In interatrial septum and atrial walls more NPY-IR than TH-IR fibers were evident, and NPY-IR, but not TH-IR, neurons were detected in intrinsic ganglia. Double-label immunohistochemistry provided further evidence that NPY is located in ventricular and atrial noradrenergic nerves, but is also located in nonnoradrenergic nerves in atria.

Aging↗
PubMed1994

Alpha-melanocyte-stimulating hormone and N-acetyl-beta-endorphin immunoreactivities are localized in the human pituitary but are not restricted to the zona intermedia.

The adult human pituitary lacks a well defined intermediate lobe, and it is uncertain whether the POMC cells that remain in the zona intermedia represent melanotropes or corticotropes. In the present study, we investigated whether the N-acetylated beta-endorphin- and alpha-MSH-related peptides that are characteristically produced by melanotropes in the rat and other species are localized in the human pituitary. Sequential gel filtration and ion exchange HPLC analysis revealed that small amounts of alpha-N-acetyl-beta-endorphin-(1-31), as well as beta-endorphin-(1-27) and beta-endorphin-(1-26), were detectable in human pituitary extracts, although beta-endorphin-(1-31) was clearly the major form. Consistent with this analysis, low levels of alpha-MSH, but not N,O-diacetyl-alpha-MSH, were identified by reverse-phase HPLC, although again, the desacetyl form of alpha-MSH predominated. Immunohistochemistry revealed that N-acetyl-beta-endorphin immunoreactivity was colocalized with ACTH and beta-endorphin in a subpopulation of zona intermedia cells. Unexpectedly, immunoreactive N-acetyl-beta-endorphin was also observed in a comparable proportion of corticotropes dispersed throughout the anterior lobe. alpha-MSH immunoreactivity was similarly distributed. These results indicate that N-acetylation is not restricted to the zona intermedia, suggesting that the strict dichotomy between corticotrope and melanotrope POMC processing observed in the rat and other species does not extend to the human pituitary.

Adrenocorticotropic Hormone↗
PubMed1993

Diminished neuropeptide Y and dopamine beta-hydroxylase immunoreactivity in a guinea pig model of left ventricular hypertrophy.

OBJECTIVE: The aims of the study were to determine the effect of chronic pressure overload of the left ventricle on the density and distribution of neuropeptide-Y-like immunoreactive (NPY-LI) nerve fibres in heart and to compare any changes to those observed in adrenergic nerve fibres, identified by dopamine beta-hydroxylase immunoreactivity. METHODS: Pressure overload was produced in female adult guinea pigs by constriction of the abdominal aorta, using a modified Weck haemoclip. The same operation was performed on a separate group of animals except that no clip was placed around the aorta. Five weeks after surgery, animals were anaesthetised, and the hearts were fixed by perfusion for immunohistochemistry. Cryostat sections were stained, using an indirect peroxidase/antiperoxidase method, for NPY or dopamine beta-hydroxylase. RESULTS: Aortic stenosis caused a 45% increase in left ventricular weight and a 58% increase in left atrial weight at 5 weeks postsurgery. Pulmonary oedema, a sign of cardiac failure, was evident in most of the animals with aortic stenosis. Immunohistochemical studies showed that in atria and right ventricles from animals with abdominal aortic stenosis the distribution and density of NPY-LI nerve fibres were similar to those in the sham operated guinea pigs. However, the left ventricles obtained from the animals with aortic stenosis were nearly devoid of NPY-LI nerve fibres. The density of dopamine beta-hydroxylase-LI nerve fibres was also substantially reduced in the hypertrophied left ventricles. CONCLUSIONS: Aortic stenosis resulting in left ventricular hypertrophy caused a nearly complete loss of NPY-LI and dopamine beta-hydroxylase-LI nerve fibres from the left ventricle. The parallel reduction in both neuropeptide Y and dopamine beta-hydroxylase is in accordance with the association of neuropeptide Y with sympathetic (adrenergic) nerve fibres in the left ventricle and suggests that chronic left ventricular hypertrophy causes a severe degeneration of sympathetic axons supplying this chamber and/or reduces the ability of these sympathetic neurones to maintain normal levels of neurotransmitter related enzymes and neuropeptides.

Animals↗
PubMed1993

Calcium regulation of intracellular pH in pituitary intermediate lobe melanotropes.

The regulatory activities of both intracellular calcium ([Ca2+]i) and intracellular pH (pHi) have greatly increased interest in the study of their interdependence. We have designed an epifluorescence video microscope that will image the fluorescence from two ratio dyes, indo-1 (for [Ca2+]i) and SNARF-1 (for pHi) at video rates. We examined primary cultures of pituitary intermediate lobe melanotropes loaded with both dyes. After experimentation, cells were positively identified by fluorescence immunohistochemistry. K(+)-induced depolarization of melanotropes produced increases in [Ca2+]i due to activation of L-type Ca channels. A secondary Ca2+ peak or oscillations were often seen. After treatment with carbonyl cyanide m-chlorophenylhydrozone, depolarization produced a rise in intracellular [Ca2+]i as well as oscillations. After thapsigargin or cyclopiazonic acid treatment, depolarization produced a primary Ca2+ elevation, but the secondary Ca2+ changes disappeared. This suggests that the oscillations were due to Ca2+ release from an endoplasmic reticulum type of intracellular store. All of these increases in [Ca2+]i were also directly coupled to a rise in intracellular H+. The close association between intracellular Ca2+ and H+ suggests that the observed pHi changes were due to the release of H+ upon binding of Ca2+ to intracellular buffers. This direct obligate coupling of intracellular Ca2+ and H+ suggests the possibility that pH-dependent cellular processes are directly activated by sudden increases in intracellular Ca2+ levels. This second messenger type of signaling system would be activated whether the Ca2+ was released from intracellular stores or entered the cell via plasma membrane Ca2+ channels.

Animals↗
PubMed1992

N-acetylation and C-terminal proteolysis of beta-endorphin in the anterior lobe of the horse pituitary.

beta-Endorphin is post-translationally processed to both N-acetylated and C-terminally shortened derivatives in the anterior lobe of the horse pituitary, a processing pattern qualitatively different from that of the rat and virtually every other mammalian species. Thus, separation of the molecular forms of beta-endorphin using gel filtration and ion exchange chromatography showed that the horse anterior lobe primarily contains beta-endorphin-1-31 and N-acetyl-beta-endorphin-1-27 along with smaller amounts of beta-lipotropin, beta-endorphin-1-27, and N-acetyl-beta-endorphin-1-31 and -1-26, in contrast to the rat anterior lobe, which contains approximately equal amounts of beta-lipotropin and beta-endorphin-1-31. Immunohistochemical experiments using an antiserum which specifically recognizes N-acetylated beta-endorphin peptides confirmed that N-acetyl-beta-endorphin immunoreactivity is present in the anterior lobe of the horse, but not the rat. The intermediate lobe of both species primarily synthesizes N-acetylated, C-terminally shortened beta-endorphin peptides, and while distinct species differences do occur, they were relatively minor, consisting of quantitative differences in the relative proportion of each peptide. These results are consistent with earlier reports that beta-endorphin processing in the rat pituitary is tissue specific; the anterior and intermediate lobes produce entirely different sets of beta-endorphin peptides. In the equine pituitary, however, both pituitary lobes produce the same multiple beta-endorphin forms, possessing both opioid and nonopioid properties, although their relative amounts differ.

Acetylation↗
PubMed1992

The onset of dopaminergic innervation during ontogeny decreases melanotrope proliferation in the intermediate lobe of the rat pituitary.

The onset of dopaminergic innervation and its effects on melanotrope proliferation were investigated in the rat pituitary intermediate lobe. Dopamine, and its synthetic rate-limiting enzyme tyrosine hydroxylase, were first detected immunohistochemically on late post-natal day 3 or early postnatal day 4. Axon density was highest at the neural lobe/intermediate lobe border, and decreased toward the pituitary cleft. By postnatal day 10, the adult pattern of tyrosine hydroxylase immunoreactivity was established and remained through post-natal day 14. Neurointermediate lobe dopamine levels, measured by HPLC, correlated well with the increased axon density observed in the immunohistochemical studies. Dopamine could not be measured by our assay (100 fg limit) until post-natal day 3 (439.32 fg/NIL). Dopamine concentration increased to 2.09 +/- 0.425 ng at PN 4, 86.31 +/- 20.42 ng at PN 7, 168.72 +/- 18.37 ng at PN 10. Melanotrope proliferation was determined by [3H]thymidine incorporation before and after innervation. Concomitant with the onset of innervation, the proliferation index dropped from 13.4 +/- 0.01% to 6.5 +/- 0.002% at PN 4, and continued to decrease until a level of 3 +/- 0.003% was established by PN 10. To confirm the inhibitory action of dopaminergic innervation on melanotrope proliferation, rat neonates were injected intracisternally with 150 mg 6-hydroxydopamine to destroy dopaminergic axons within the intermediate lobe. Measurement of dopamine concentrations in neurointermediate lobes of injected animals showed a decrease in dopamine levels as compared to controls. From PN 4 (0.88 +/- 0.165 ng), DA levels gradually increased during development: at PN 5, [DA] = 0.689 +/- 0.104 ng; PN 6 [DA] = 11.60 +/- 2.24 ng; PN 7 [DA] = 20.93 +/- 3.80 ng; and PN 10 [DA] = 27.95 +/- 3.46 ng. Melanotrope proliferation also increased in 6-hydroxydopamine-treated animals. At PN 4, the onset of innervation reduced the pre-innervation proliferation index to 8.75 +/- 0.002%, only a 30% reduction in contrast to the greater than 50% decrease observed in control animals. A stable proliferation level of approximately 7.5% persisted in all subsequent stages with 6-OHDA administration. Our results demonstrated the time of dopamine innervation onset and a characteristic developmental pattern for axons within the rat intermediate lobe. The onset of innervation and increased dopamine concentration suggests increased dopaminergic control of the melanotropes, illustrated specifically by a decrease in their level of proliferation. This is the first presentation of evidence showing that dopaminergic innervation within the intermediate lobe of the rat pituitary regulates melanotrope proliferation.

Animals↗
PubMed1988

Neuropeptide Y in the rat nucleus accumbens: ultrastructural localization in aspiny neurons receiving synaptic input from GABAergic terminals.

The ultrastructure, afferent input, and sites of termination of neurons containing neuropeptide Y-like immunoreactivity (NPY-LI) were examined in the adult rat nucleus accumbens by using the peroxidase-antiperoxidase (PAP) method. The NPY-LI was seen in sparsely distributed, spindle-shaped perikarya having cross-sectional diameters of 15-20 microns. These perikarya exhibited highly invaginated nuclear membranes and thin rims of cytoplasm containing Golgi lamellae, dense-core vesicles, and other organelles. A few large, principally aspiny, dendrites also showed NPY-LI. The dendrites received synaptic input from unlabeled terminals forming both symmetric and asymmetric junctions. Immunolabeling for NPY was evident in other processes that were not clearly differentiated as dendrites or axons. These were seen primarily near glial processes and the basal laminae of blood vessels. A few myelinated and many unmyelinated axons and axon terminals also were labeled for NPY. These terminals contained numerous, small (40-60 nm), clear and one or more large (80-100 nm) dense core vesicles. Forty-seven percent (27 out of 57) of the terminals containing NPY-LI formed symmetric junctions with unlabeled dendrites or dendritic spines. The remainder lacked recognizable densities within single planes of section. The neurons exhibiting NPY-LI in the nucleus accumbens were characterized further with respect to their afferent input from terminals labeled for the GABA-synthesizing enzyme, glutamic acid decarboxylase (GAD). Immunogold labeling of a rabbit antiserum against NPY and PAP labeling for a sheep antiserum to GAD were sequentially applied to the same sections. The GAD-labeled terminals formed symmetric junctions primarily with the more numerous unlabeled dendrites. However, a few synaptic junctions also were detected between the GAD-labeled terminals and dendrites showing immunogold labeling for NPY. We conclude (1) that in the rat nucleus accumbens, NPY-LI is found principally in neurons of the aspiny type and (2) that the output from these presumably intrinsic neurons to other neighboring neurons or blood vessels is at least partially modulated by GABA.

Animals↗
PubMed1988

Nuclear protein kinase substrates in AtT-20 cells: translocation of a 14 kDa phosphoprotein.

In this work we characterize some acidic nuclear substrates of protein kinase C (PKC) and cyclic AMP-dependent protein kinase (PKA), using intact anterior pituitary corticotrophic tumor cells (AtT-20/D16-16). It was found that, as in the cytosolic fraction, substrates for both PKC and PKA exist in the nucleus and that changes in the phosphorylation states of a few of these phosphoproteins are mediated by both kinases. One of the phosphoproteins examined, a 14 kDa phosphoprotein (pp14) described previously, exhibited a phorbol-ester induced translocation from nucleus to cytosol in pulse-chase experiments utilizing 35S-methionine labeling. These results suggest that pp14 may be involved in signal transduction in AtT-20 cells. Although its identity remains to be determined, a 14 kDa DNA-binding protein was also seen in nuclear extracts of AtT-20 cells.

Animals↗