LHRH-like immunoreactive material was separated from human placental homogenates by chromatography and tested against two antibodies directed respectively against the C- or the N-terminus (N- and C-antibodies) of the synthetic peptide. Extraction yield was 94 per cent as assessed by the recovery of the radiolabelled LHRH added to the homogenates. Placental LHRH-like immunoreactivity in crude extracts appeared largely overestimated, due to interference of endogenous substances non-related to LHRH. After elimination of this contamination, a higher residual activity was detected with the C- than with the N-antibodies. Eluates tested under these conditions contained only 550 pg of LHRH-like material per kg of placenta. This concentration was comparable whether placental tissue had been sampled at 12 weeks of pregnancy or at the end of gestation. The corresponding peptides however neither co-eluted with synthetic LHRH nor with catabolite fragments of the peptide. It is concluded that the human placenta contains a lesser amount of LHRH-like material than anticipated on the basis of earlier results and that this material does not correspond to the native decapeptide.

Antibody Specificity↗

PubMedDate not supplied

Interference of endogenous B endorphin with opiate binding in the anterior pituitary.

Opiates have been shown to affect prolactin secretion directly at the pituitary level. However, opiate binding sites have not been characterized on anterior pituitary tissue. In the present work we show that a soluble factor present in anterior pituitary membrane preparation inhibit opiate binding on striatal membranes. The high concentrations of B-endorphin like immunoreactivity recovered from the supernatant of pituitary membrane preparations seem to account for this effect on striatal binding and to be responsible for a partial masking of pituitary opiate receptors. We thus attempted to improve membrane fractionation procedure in order to decrease B-endorphin contamination and thus to increase binding of opiate ligands on pituitary membranes. 3-H Etorphine binds to pituitary membranes with a constant of association (kon) of 20.8 10(6) M-1 min-1 and a constant of dissociation (hoff) of 33 min-1. Saturation experiments with 3-H Etorphine and 3-H ethylketocyclazocine indicate the presence of a single population of specific binding sites (KD) value of 12.2 +/- 3.9 nM a maximal binding capacity of 55.9 +/- 10.7 fm/mg protein for 3-H Etorphine binding). These results demonstrate the presence of opiate binding sites in the anterior pituitary. However the difficulty of the membrane preparation and the low density of binding sites did not allow a complete pharmacological characterization of these sites.

Animals↗

PubMedDate not supplied

Autoradiographic localization of a non-reducible somatostatin analog (125I-CGP 23996) binding sites in the rat brain: comparison with membrane binding.

The regional distribution of somatostatin binding sites in the rat brain was determined by quantitative autoradiography, using 125I-CGP 23996, a non-reducible somatostatin analog. In preliminary experiments, kinetic properties of 125I-CGP 23996 binding to rat brain membranes and slide mounted frozen brain sections were compared and found similar. In addition, distribution of 125I-CGP 23996 and 125I-N-Tyr-SRIF14 binding sites on membrane prepared from 10 different rat brain structures were closely correlated (r = 0.91, 2 p less than 0.01), indicating that the non-reducible analog recognizes the same binding site as the Tyr-extended native peptide. Highest levels of 125I-CGP 23996 binding sites were found in anterior temporal, frontal and cingular cortex as well as hippocampus. Moderate levels were found in the remaining part of the limbic system including amygdala, olfactory tubercles and bed nucleus of the stria terminalis. In the brain stem, nuclei involved in the auditory system such as the ventral cochlear nucleus and the superior olive nucleus, contained high levels of 125I-CGP 23996 binding sites. The distribution of 125I-CGP 23996 binding sites roughly correlated with that of the endogenous peptide in most structures, except in the mediobasal hypothalamus.

Amygdala↗

PubMedDate not supplied

Vasoactive intestinal peptide induces a transient release of growth hormone in the rat.

Effects of vasoactive intestinal peptide (VIP) on growth hormone (GH) secretion were investigated both in vivo on freely moving, intact or mediobasal hypothalamic lesioned rats, and in vitro in incubation or superfusion systems of anterior pituitary tissue. In vivo, IV injection of VIP induced a rapid but transient increase in plasma GH levels in all animals and in vitro, VIP stimulated GH release from incubated or superfused rat pituitaries in a concentration dependent manner. This increase was potentiated by forskolin, a potent activator of adenylate cyclase. These findings indicate that VIP exerts a direct stimulating action on somatotrophs and that the effect seems to imply a coupling of VIP receptors with cAMP production.

Animals↗

PubMedDate not supplied

Evidence for a single class of somatostatin receptors in ground squirrel cerebral cortex.

In the present study we characterized high-affinity somatostatin (SRIF) binding sites (Kd = 2.06 +/- 0.32 nM and Bmax = 295 +/- 28 fmol/mg protein) in cerebral cortex membrane preparations of European ground squirrel using 125I-[Tyr0-D-Trp8]-SRIF14 as a radioligand. The inhibition of radioligand specific binding by SRIF14, as well as by its agonists (SRIF28, Tyr0-D-Trp8-SRIF14, SMS 201 995) was complete and monophasic, thus revealing a single population of somatostatinergic binding sites. Radioautographic analysis of 125I-[Tyr0-D-Trp8]-SRIF14 labeled brain sections confirmed the results of our biochemical study. The homogeneity of SRIF binding sites in the ground squirrel neocortex was not dependent on the animal's life-cycle phase.

Animals↗

PubMedDate not supplied

125I-[Tyr0,D-Trp8]somatostatin-14 binding sites in the locus coeruleus of the rat are located on both ascending and descending projecting noradrenergic cells.

Radioautographic determinations of 125I-[Tyr0,D-Trp8]somatostatin-14 (125I-SRIF) binding sites were performed on frozen serial sections of the locus coeruleus (LC) of control rats and of rats subjected to either bilateral microinjections of 6 hydroxydopamine (6-OHDA) into the LC or unilateral microinjection into the ascending noradrenergic bundles. These experiments were performed in order to determine whether 125I-SRIF binding was localized to noradrenergic-containing cells and in which regions the cells which contain the binding sites are projecting. The extent of the lesions was assessed by measuring norepinephrine (NE) levels in the hippocampus (88% decrease as compared to sham-operated animals) for bilateral LC lesions and in the frontal cortex (87% reduction vs. contralateral side) for unilateral bundle lesions. In control rats, 125I-SRIF binding sites were restricted to the boundaries of the LC and followed closely the distribution of tyrosine hydroxylase-labeled cells. Three weeks after bilateral injections of 6-OHDA, 125I-SRIF binding decreased by 79% in all regions of the LC. In contrast, unilateral destruction of the ascending noradrenergic bundles resulted in a moderate decrease only in the middle part of the LC with a more important effect in the dorsal (55%) than in the ventral (24%) portion of the nucleus. These data demonstrate that: 1) most SRIF receptors in the LC are located in the vicinity of NE-containing cell bodies and 2) NE-containing cells bearing SRIF receptors project to the forebrain as well as to other terminal areas located more caudally in the brain. These data suggest a general role for SRIF in the control of the multiple functions of the LC.

Animals↗

PubMedDate not supplied

Choline acetyltransferase and somatostatin levels in aged Microcebus murinus brain.

beta-Amyloid protein (beta-AP) deposits, analoguous to those found in Alzheimer's disease (AD) are observed in the brain of aging Microcebus murinus. Because choline acetyltransferase (ChAT) activity and somatostatin (SRIH) content are consistently decreased in AD, we tested whether such changes could be observed in middle aged to aged Microcebus cerebral cortex and whether they were accompanied by beta-AP deposits. A positive correlation was observed between age and ChAT activity. By HPLC, SRIH immunoreactivity eluted as four peaks, two of which being identical with SRIH-28 and SRIH-14 while the other two likely represented precursor forms. Cortical SRIH content was not significantly affected by age. ChAT activity and SRIH content were not significantly correlated. Amyloid angiopathy was observed in every brain examined and the presence of cortical lesions analoguous to senile plaques observed in the oldest case only which did not demonstrate important alterations in ChAT and somatostatin levels.

Aging↗

PubMedDate not supplied

Participation of serotonin-containing neurons in the suckling-induced rise in plasma prolactin levels in lactating rats.

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5-Hydroxytryptophan↗

PubMedDate not supplied

Neural and pituitary mechanisms involved in growth hormone regulation.

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Animals↗

PubMedDate not supplied

ADP-ribosylation of G alpha i and G alpha o in pituitary cells enhances their recognition by antibodies directed against their carboxyl termini.

Using antibodies raised against synthetic peptides of heterotrimeric GTP binding proteins, we demonstrate the presence of G alpha s, G alpha i1,2, G alpha i3, G alpha o2, and G beta subunits in pituitary cells. Pretreatment of pituitary cells with cholera toxin diminished the immunoreactivity of G alpha s and this decrease was kinetically coupled to the rate of G alpha s ADP-ribosylation. ADP-ribosylation by islet activating protein (IAP or Bordetella pertussis toxin) of G alpha i and G alpha o enhanced their immunoreactivities to antibodies raised against synthetic decapeptides that correspond to the G alpha carboxyl termini. Such enhancement was not observed when antibodies directed against the NH2-termini were used. These findings are consistent with the fact that ADP-ribosylation by IAP occurs on the cysteine located in the carboxyl terminal part of G alpha i and G alpha o. These observations mean that the kinetics and extent of Gi and Go ADP-ribosylation by IAP in whole pituitary cells and membrane preparations can be followed. It could be that ADP-ribosylation causes conformational changes in G alpha i and G alpha o. Indeed, we observed that ADP-ribosylated G alpha i was more sensitive to trypsin proteolysis and that the ADP-ribosylation rates of G alpha i and G alpha o in whole cells were comparable to the rate of loss of coupling between inhibitory neurohormone receptors and adenylyl cyclase.

Adenosine Diphosphate Ribose↗