IDENTIFICATION OF A RAT GHRH-LIKE SUBSTANCE AND ITS MESSENGER RNA IN RAT TESTIS

Abstract

The adult testis is a source of hypothalamic neuropeptides including TRH, GnRH, POMC and CRF. The role of these neuropeptides as paracrine regulators of gonadal function in the testis remains unknown. We postulated that growth hormone releasing hormone (GHRH) might also be measureable in rat testis. Testes from ten post pubertal rats (400-425g, .apprx.90d) were extracted and assayed in an enzyme linked immunosorbent assay (ELISA) which utilized an anti-rat GHRH antiserum and had a sensitivity of 30 pg. We detected 1.62 .+-. 0.17 (M .+-. SEM) ng of GHRH-like substance per g testis. The GHRH content of hypothalamus from these animals was 2.70 .+-. 0.24 ng/g (N = 7,6 hypothalami/N). Fat served as a negative control tissue in which no GHRH-like material was detectable. To confirm the finding and in attempt to determine if it represented local production of GHRH, we measured GHRH mRNA using a cRNA probe subcloned from a cDNA for rat GHRH. Abundant levels of hybridizing RNA were found in mature testis by dot-blot hybridization (319% .+-. 30% of the mean hypothalamus value). Northern analysis revealed that the rat hypothalamic GHRH mRNA was 715 nucleotides. In contrast, testicular GHRH(-like) mRNA was 1750 nucleotides. We conclude that there are high levels of a GHRH-like substance in mature rat testis which is present both at the level of protein product and gene transcript. The mRNA for this GHRH-like substance is substantially larger than the GHRH transcript from hypothalamus.