A radioimmunoassay specific for [Gln8]LH-RH: application in the confirmation of the structure of chicken hypothalamic luteinizing hormone-releasing hormone.

In the first report on the chemical structure of a nonmammalian LH-RH, chicken hypothalamic LH-RH was demonstrated to be [Gln8]LH-RH [2-4]. However, these studies and subsequent reports [7,8] did not totally exclude the possibility of a reverse sequence of the two amino acids Leu-Gln. In view of the recently described structure of salmon brain LH-RH as [Trp7,Leu8]LH-RH [9], we undertook to confirm our earlier conclusion that chicken LH-RH is [Gln8]LH-RH and not [Gln7, Leu8]LH-RH. The immunologic, chromatographic and biological properties of natural chicken hypothalamic LH-RH were compared with those of the two synthetic peptides, [Gln8]LH-RH and [Gln7,Leu8]LH-RH. A radioimmunoassay highly specific for [Gln8]LH-RH was developed. Natural chicken LH-RH cross-reacted fully with the antiserum which requires the COOH-terminal Gln8 to Gly10-NH2 for binding, while [Gln7,Leu8]LH-RH showed less than 0.1% cross-reaction. On a high resolution reverse phase high performance liquid chromatography system, natural chicken LH-RH co-eluted with [Gln8]LH-RH and was well separated from [Gln7,Leu8]LH-RH. In a chicken anterior pituitary cell bioassay, natural chicken LH-RH and [Gln8]LH-RH were equipotent in stimulating luteinizing hormone release, while the relative potency of [Gln7,Leu8]LH-RH was 4.4%. These data, in particular the use of a specific [Gln8]LH-RH antiserum, provide conclusive evidence that chicken LH-RH is [Gln8]LH-RH.

Ihibition of rat brain norepinephrine N-methyltransferase in vitro and in vivo by chloro-substituted 1-aminoindans.

Norepinephrine N-methyltransferase from rat brain stem was inhibited in vitro by 1-aminoindans with chlorine substituents on the aromatic ring. The order of inhibitory potency among these compounds was 4.5-dichloro greater than 4-chloro greater than 5-chloro approximately or equal to 5,6-dichloro approximately or equal to 6,7-dichloro greater than 6-chloro greater than 7-chloro. All except the 7-chloro compound were more potent inhibitors than the parent unsubstituted 1-aminoindan. 4,5-Dichloro-1-aminoindan was a competitive inhibitor in kinetic studies with L-norepinephrine as the variable substrate; its Ki value determined from a Dixon plot was 3.3 x 10(-7) M. At doses of 10-40 mg/kg i.p., this compound inhibited brain stem and hypothalamic norepinephrine N-methyltransferase in vitro and lowered hypothalamic concentrations of epinephrine in rats, effects that lasted for several hours. 4,5-Dichloro-1-aminoindan may be a useful pharmacologic tool for studies of epinephrine-forming neurons in brain.