Regulation of calcitonin gene-related peptide expression in dorsal root ganglia of rats by female sex steroid hormones

Calcitonin gene-related peptide (CGRP), a potent vasodilator primarily synthesized in dorsal root ganglia (DRG) neurons, has been shown to decrease vascular resistance and thus regulate blood flow to a variety of organs in rats. Serum CGRP levels in the human have been reported to increase with pregnancy and decrease postpartum. It has been suggested that female sex steroid hormones play a role in cardiovascular function, but the mechanisms are unknown. In this study, we examined the effects of estradiol-17 beta (E-2) and progesterone (P-4) on the expression of CGRP in DRG in adult rats both in vivo and in vitro. Ovariectomized (ovx) animals were injected s.c. with 5 mu g E-2, 4 mg P-4, or 5.01 mu g E-2 + 4 mg P-4 in 0.5 ml sesame oil or with oil only, and groups of 4 rats were killed at 0, 24, or 48 h. DRGs were then removed and analyzed for CGRP mRNA and immunoreactive (i-)CGRP content by Northern blotting and RIA, respectively. Primary cultures of DRG neurons from adult female rats were used to assess the effects of varying doses of E-2 (1, 10, 100 nM), P-4 (10, 100, 1000 nM), or E-2 (10 nM) + P-4 (100 nM) in the absence or presence of nerve growth factor (NGF; 20 ng/ ml); and CGRP mRNA content in the cells and i-CGRP in the medium were quantitated at 24 or 48 h after incubation. Results of in vivo studies showed that E-2 caused a significant increase in CGRP mRNA at 24 h (1.8-fold) and in i-CGRP levels both at 24 h (2.8-fold) and at 48 h (3.4-fold) in DRG of ovx rats. P-4 also stimulated expression of both CGRP mRNA and i-CGRP. In the in vitro studies, either E-2 or P-4 alone or the two in combination were without effect on CGRP expression in cultured DRG neurons at all the doses tested. However, in the presence of NGF, both CGRP mRNA and peptide levels were significantly enhanced by E-2, P-4, and E-2 + P-4 in a time-dependent (2.0- to 2.8- fold at 24 h, 3.0- to 5.0-fold at 48 h) and dose-dependent manner, with maximal effects achieved at 1.0 nM (E-2) and 100 nM (P-4) at 24 h of incubation. In summary, both E-2 and P-4, either alone or in combination, stimulate CGRP peptide synthesis in DRG neurons through increasing CGRP mRNA. The effects of these steroid hormones are mediated through amplifying the NGF-induced synthesis of CGRP in these neurons. Thus, we propose that the cardiovascular functions of female sex steroid hormones may be mediated, at least in part, by the up-regulation of neuronal CGRP synthesis, via NGF-mediated mechanisms.