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and Luteinizing Hormoneβ Messenger Ribonucleic Acid (RNA) of Male and Female Rats after Castration: Quantitation Using an Optimized RNA Dot Blot Hybridization Assay^*

Department of Internal Medicine, Division of Endocrinology and Metabolism, Department of Pharmacology and the Reproductive Endocrinology Program, University of Michigan Ann Arbor, Michigan 48109

Address requests for reprints to: J. C. Marshall, Department of Internal Medicine, Division of Endocrinology and Metabolism, University of Michigan Hospital, Ann Arbor, Michigan 48109.

Abstract

In this study we examined the changes in a and LHβ mRNAs in anterior pituitaries of male and female rats after castration. mRNA concentrations were measured by an optimized RNA dot blot hybridization assay. Rat a and LHβ cDNAs were nick-translated to specific activities of 2–5 x 108 cpm/µg and were used as hybridization probes. The total RNA per assay, RNA per dot, and saturating amounts of probe were optimized. The intra- and interassay coefficients of variation were 5% and 28%, respectively.

Both and LHβ mRNA concentrations increased after castration, but marked differences were observed in the kinetics of responses in male and female rats. In males, a and LHβ mRNAs were increased by 24 h postcastration (by 25% and 38%, respectively), and 4- to 5-fold increases over intact controls were evident by 18 days, mRNA rose rapidly and had doubled by 2 days, whereas LHβ mRNA concentrations showed a similar increase by 6–7 days postcastration. The slower rise in LHβ mRNA was associated with a transient decline in serum and pituitary LH concentrations between 2 and 6 days after castration.

In female rats, mRNA increased more slowly, concentrations had doubled by 10 days, while a similar increase in LHβ mRNA occurred 7 days after castration. Thereafter, both subunit mRNAs continued to rise, and by day 20 mRNA was increased 5-fold and LHβ mRNA 16-fold over values in intact females. Serum and pituitary LH concentrations rose gradually, and both were increased by 7–10 days after castration. The increase in serum and pituitary LH followed a time course similar to that of the progressive rise in LHβ mRNA concentrations.

These data show that an increase in steady state LH subunit mRNA concentrations is one of the mechanisms involved in increased gonadotropin biosynthesis and secretion after castration. The kinetics of LH subunit mRNA and LH secretory responses are different in male and female rats and suggest that the concentration of LHβ mRNA may be a limiting factor in LH secretion. (Endocrinology 119: 691–698, 1986)

Footnotes

^* This work was supported by USPHS Grant HDl 1489 (to J. C. M.).

Received January 2, 1986.

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