NEUROTENSIN SYNTHESIS IN ARCUATE NEUROSECRETORY CELLS

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The broad objective of this proposal is to clarify the function of neurotensin (NT) synthesis and-release by neurosecretory cells located in the dorsomedial division of the hypothalamic arcuate nucleus in the rat. Regulation of NT synthesis in this region follows a pattern that implies NT involvement in female-specific, estrogen-dependent secretion of one or more adenohypophysial hormones, particularly prolactin and the gonadotropins. To test this hypothesis, NT antiserum will be used to selectively interfere with endogenously released NT in conscious, freely moving females at proestrus, and the effect of this intervention on circulating levels of prolactin, luteinizing hormone, and follicle- stimulating hormone will be determined. A primary aim of this proposal is to determine in the female the extent to which estrogen-inducible synthesis of NT occurs in tuberoinfundibular dopaminergic neurons under physiological conditions. To that end, a double-label method of in situ hybridization will be used to determine the number and distribution of dorsomedial arcuate neurons that coexpress mRNA encoding NT and neuromedin N (NT/N mRNA) and tyrosine hydroxylase mRNA under different endocrine conditions. The same technique will also be used to investigate likely coexpression of NT/N mRNA with either estrogen receptor mRNA or NT receptor mRNA in this region and thus to identify NT neurons subject to direct regulation by estrogen or NT itself. A basic premise of these studies is that heterogeneous coexpression of the NT/N gene with certain other genes in dorsomedial neurons will reveal functional subgroups of NT-synthesizing neurons in this region. Hormonal regulation of NT/N mRNA expression in these neurons will be investigated to obtain additional clues to their function. In situ hybridization will be used to assess the possibility that prolactin or progesterone influence NT/N mRNA expression in the dorsomedial arcuate nucleus. Also, in situ hybridization with an intron-specific probe will be used to determine whether estrogen increases NT/N mRNA levels through increased transcription of the NT/N gene. The time course over which estrogen increases levels of NT/N primary transcript and mature NT/N mRNA will indicate whether estrogen- induced synthesis of NT could plausibly mediate specific neuroendocrine events. Steady-state levels of NT and neuromedin N will be measured to determine whether estrogen alters their ratio in the median eminence or neurointermediate lobe. Finally, in situ hybridization will be used to determine whether adult sex differences in plasma estrogen levels account for sexually differentiated expression of NT/N mRNA in dorsomedial arcuate neurons. By clarifying the role of NT in normal neuroendocrine function, these studies will provide a basis for future investigation of NT involvement in human neuroendocrine disorders.