Thyroliberin acetate salt

Binding of TRH to specific cell surface receptors on clonal GH4C1 cells is followed within 10 min by receptor sequestration and over 24 h by receptor down-regulation. These experiments were designed to determine if TRH-activated second messenger systems are responsible for changes in receptor localization or number. BAY K8644 and A23187, which increase intracellular calcium, alone or together with 12-O-tetradecanoyl phorbol acetate (TPA), which activates protein kinase C, did not appear to internalize TRH receptors. Drug treatment did not alter the rate of [3H]MeTRH association or internalization, determined by resistance to an acid/salt wash, or the amount of [3H]MeTRH able to bind at 0 C, where only surface receptors are accessible. TPA (0-100 nM) alone or in combination with BAY K8644 or A23187, also failed to change receptor number or affinity after 48 h when TRH caused a 75% decrease in the density of specific binding sites. Chlordiazepoxide has been reported antagonize TRH binding and TRH-induced phospholipid breakdown. Chlordiazepoxide shifted the dose-response curves for TRH stimulation of PRL release and synthesis to the right, and did not change PRL release alone. The affinity of receptors for chlordiazepoxide was not affected by a nonhydrolyzable analog of GTP whereas affinity for TRH was decreased; these properties are consistent with the classification of chlordiazepoxide as a competitive antagonist. Several experiments tested whether chlordiazepoxide would cause receptor internalization and down-regulation. Chlordiazepoxide did not appear to internalize TRH receptors, because TRH-binding sites became available rapidly and at the same rate after they had been saturated with chlordiazepoxide at 0 or 37 C.(ABSTRACT TRUNCATED AT 250 WORDS)

This paper describes the implementation of an ultraviolet (UV) laser (Spectra Physics 171-18 with 3 lines: 334, 351 and 364 nm in UV) as light source for fluorescence confocal scanning microscopy. With this instrument it is possible to use fluorophores not previously available for confocal laser microscopical imaging of fluorophores such as fluoro-gold and AMCA. In the study we show confocal laser microscopical imaging of fluorescent motoneurons labelled by retrograde transport of fluoro-gold and AMCA-fluorescent axon terminals labelled with antisera against immunogenes as thyrotropin-releasing hormone (TRH) and calcitonin gene-related peptide (CGRP). These two fluorophores may be recorded simultaneously or separately by using a filter that suppresses the emission of one of the fluorophores. The described instrument should also be useful in applications involving detection of monoamines by the Falck-Hillarp technique, as well as measurements of cytosolic free calcium by indicators such as Fura-2 and Indo-1. Measurements performed in reflected and fluorescence light indicated that the resolution along the optical axis improved by about 25% when UV (351 nm) is used instead of visible light (514 nm). This figure is close to that expected on theoretical basis. There are, however, also serious problems related to the use of UV excitation. Firstly, objectives must be selected based on their UV transmission properties. Secondly, chromatic aberration may cause a substantial focal shift between illuminating and emitted light, calling for a flexible instrumental design in order to allow for compensation. As shown here, this problem can be circumvented by using reflecting objectives but at a price of lower resolution compared with high-aperture refracting objectives.