Fate of Thyrotropin Releasing Hormone after Binding and Stimulation of Prolactin Release by GH3 Cells. Evidence for Release of Unmodified [&lt;sup&gt;3&lt;/sup&gt;H] TRH

Gourdji, D.; Tixier‐Vidal, A.; Levine‐Pinto, H.; Pradelles, P.; Morgat, Jean‐Louis; Fromageot, P.

doi:10.1159/000122484

Cited by 11 publications

(6 citation statements)

References 2 publications

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“…Indeed previous analysis by thin layer chromatography of tritiated material associated to intact cells exposed for increasing durations to ('H)TRH revealed a very slow rate of degradation: 5% after 2 h. 30% after 20 h. In addition, the radioactive material which remains associated to the cells after I h of postincu bation at 37 °C following ('H)TRH uptake for 30 min to 20 h consisted mostly of TRH (91 and 82%, respectively) ■ [13]. Moreover, the pituitary enzymes which degrade TRH have an optimal pH of activity at 7.4 and are localized both in particulate and soluble fractions [1], There are thus very few chances that they may be affected by chloroquine.…”

Section: Discussionmentioning

confidence: 85%

“…Using autora diography of 1 -¡im thick sections of araldite-embedded cells as well as fractionation methods, we have previously shown that in these conditions TRH is internalized up to the nu cleus without being degraded [3,13]. Moreover, when intact GH3 cells previously loaded at 37 °C with (3H)TRH and thoroughly washed on ice were postincubated at 37 °C, un modified (3H)TRH dissociated from the cell with a half time value of 20 min [10,13]. At present, the mechanisms which are involved in TRH internalization and release are, nevertheless, completely unknown.…”

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confidence: 99%

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Effect of Chloroquine on Thyroliberin Interaction with Clonal Rat Prolactin Cells

Tixier-Vidal¹,

Moreau²,

Picart³

et al. 1982

Neuroendocrinology

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The lysosomotropic agent, chloroquine, has been used to investigate the implication of lysosomal activity and membrane traffic in TRH binding, TRH internalization and TRH-induced stimulation of prolactin secretion in a rat prolactin cell line (GH3/B6). Chloroquine by itself does not affect cell number, cell protein and basal prolactin secretion. It does not alter TRH binding and internalization as well as both effects of TRH on the stimulation of prolactin secretion, i.e., prolactin release and prolactin production. In contrast, chloroquine partially inhibits the spontaneous dissociation of (³H)TRH from cells previously loaded with (³H)TRH and reduces prolactin release following TRH withdrawal. In addition the kinetic pattern of the TRH dissociation is modified in a manner which suggests that TRH is bound to different intracellular compartments. Chloroquine, nevertheless, does not alter the TRH-induced down regulation of (³H)TRH binding sites. Electron microscopic observations and acid phosphatases localization reveal that chloroquine elicits a disorganization of the Golgi zone and accumulation of membrane whorls within large vacuoles. This suggests that the effects of chloroquine on TRH interaction with GH3 cells may be mediated by an inhibition of membrane recycling.

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Section: Discussionmentioning

confidence: 85%

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confidence: 99%

Effect of Chloroquine on Thyroliberin Interaction with Clonal Rat Prolactin Cells

Tixier-Vidal¹,

Moreau²,

Picart³

et al. 1982

Neuroendocrinology

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“…The data concerning the analysis of tritiated material associated to intact cells (8) indicated that early biological response was induced by intact TRF. The existence of a continuous flow of TRF molecules on both sides of plasma membrane was suggested.…”

Section: 5) the Penetration Of Unmodified 3h-trf In Intact Cells Wasmentioning

confidence: 99%

“…2 ) confirms the presence of tritiated material (3) in the cell at both cytoplasmic and nuclear levels, without specific concentration on the plasma membrane. The tritiated material was identified as 'H-TRF (8) in the same experimental conditions. Contrary to the previous experiments reported (3) the relatively short time of exposure allowed here a very low background and an excellent quality of the autoradiographs.…”

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confidence: 99%

Specific tritium labelling of thyroliberin on histidyl and prolyl residues

Levine‐Pinto¹,

Morgat²,

Fromageot³

1982

Labelled Comp Radiopharmac

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SUMMARY T.ritiation of thyroliberin (L-CGlu-L-His-L-Pro-NHZ) 5starting from the L -[ A -Pro']-TRF, on prolyl and histidyl residues is being dealt with below.By catalytic tritiation of the double bond, followed by iodination of histidyl residue and catalytic deshalogenation in presence of tritium gas, TRF was labelled with a specific radioactivity close to the theoretical value. 3H-[ 2 , 5-His2-3 ,4-Pro 1 -TRF retained all biological potencies. 3Keg words : histidyl -prolyl -thyroliberin -tritiationt r i t ium .

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“…The radioactivity released into the medium was intact [3H]TRH as shown by t.l.c. [16]. In addition, the recycled material had the binding-affinity charac-Abbreviations used: HBSS/Ca2+, Hepes-buffered salt solution plus 0.5 mM Ca2+; MeTRH, 3-methyl-his-TRH; NTA, nitrilotriacetic acid; RL, receptor-4igand complex; Tf, transferrin; TRH, thyrotropin-releasing hormone; TRHR, thyrotropin-releasing hormone receptor.…”

Section: Introductionmentioning

confidence: 99%

Evidence that the thyrotropin-releasing hormone receptor and its ligand are recycled dissociated from each other

Petrou

Tashjian

1995

Biochemical Journal

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We have examined the trafficking of the thyrotropin-releasing hormone receptor (TRHR) and its ligand, after TRHR-TRH internalization in rat pituitary GH4C1 cells. After rapid ligand-induced receptor sequestration, the cell surface receptor pool was replenished. Replenishment was insensitive to inhibition of protein synthesis and was dependent on the duration of internalization; therefore, the replenished receptors were not newly synthesized but recycled. The total amount of recycled receptors decreased with increasing internalization time, resulting in only partial replenishment of the cell-surface receptor pool after prolonged incubation with ligand. Thus, in addition to a receptor recycling pathway, a non-cycling route exists for TRHR sorting; this route became dominant with increasing internalization periods. TRHR entry into these pathways was not determined by the affinity of the receptor-ligand interaction, because the extent of receptor recycling was similar after TRH- and methyl-TRH (MeTRH)-induced internalization. Unlike results with the TRHR, the TRH recycling pool was not depleted by the noncycling pathway. After multiple rounds of [3H]MeTRH internalization, the amount of cell-associated radioactivity increased with increasing internalization time due to accumulation of the ligand or its metabolites in a non-cycling pathway, but the absolute amount of recycled ligand remained constant after short or long internalization times. The difference in the proportion of TRHR and MeTRH that were diverted into a noncycling pathway indicated intracellular dissociation of the internalized TRHR-TRH complex. Dissociation of the internalized TRHR-TRH complex was dependent on the acidic pH in an intracellular compartment. Although extracellular acidic pH did not enhance cell-surface receptor-ligand (RL) dissociation, bafilomycin A1 inhibited both receptor and ligand recycling. We conclude that the TRHR-TRH system is unique among recycling receptors because, after RL sequestration, the TRHR-TRH complex becomes dissociated intracellularly via a bafilomycin A1-sensitive, acidic pH-dependent mechanism, and both the unoccupied TRHR and TRH recycle disassociated from each other.

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Fate of Thyrotropin Releasing Hormone after Binding and Stimulation of Prolactin Release by GH3 Cells. Evidence for Release of Unmodified [<sup>3</sup>H] TRH

Cited by 11 publications

References 2 publications

Effect of Chloroquine on Thyroliberin Interaction with Clonal Rat Prolactin Cells

Effect of Chloroquine on Thyroliberin Interaction with Clonal Rat Prolactin Cells

Specific tritium labelling of thyroliberin on histidyl and prolyl residues

Evidence that the thyrotropin-releasing hormone receptor and its ligand are recycled dissociated from each other

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