LHRH-I RELEASE FROM QUAIL HYPOTHALAMIC SLICES MEASURED BY SPECIFIC EIA

A competitive enzyme immunoassay (EIA) was developed for luteinizing hormone-releasing hormone (LHRH), which measures either mammalian luteinizing hormone-releasing hormone or chicken LHRH-I (cLHRH-I). There is negligible cross-reactivity with chicken LHRH-II. Assay sensitivity is 1 pg/ml and the intra- and interassay Variation are 3.4 and 10.0%, respectively. The assay was validated for measuring cLHRH-I by parallelism and quantitative recovery. Using this EIA, cLHRH-I content was measured in microdissected samples of median eminence from mature quail and chickens. Mean cLHRH-I concentrations were 1.25 +/- 0.35 and 2.10 +/- 0.25 ng/mg protein in quail and chickens, respectively. In vitro release of cLHRH-I was studied by perifusion of quail medial basal hypothalamus-preoptic area (MBH-POA) slices. Challenge with increasing concentrations of K+ resulted in significant (P < 0.05) release of cLHRH-I. The release of cLHRH-I from MBH-POA slices was also measured in response to norepinephrine (NE), epinephrine (E), and isoproterenol (ISO). Chicken LHRH-I was released in a dose-dependent manner; maximal release occurred with 10(-7) M NE, 10(-8) M E, and 10(-7) M ISO. Tissue response was optimal for experimental manipulation 6-25 hr postcollection; thereafter, the response deteriorated until 60 hr postcollection. These data extend previous studies in birds by detailed characterization of responses to neurochemical challenge and description of optimal parameters for tissue response during perifusion. (C) 1994 Academic Press, Inc.