Regulation of the cGMP-cPKG pathway and large-conductance Ca ³⁺-activated K⁺ channels in uterine arteries during the ovine ovarian cycle

The follicular phase of the ovine ovarian cycle demonstrates parallel increases in ovarian estrogens and uterine blood flow (UBF). Although estrogen and nitric oxide contribute to the rise in UBF, the signaling pathway remains unclear. We examined the relationship between the rise in UBF during the ovarian cycle of nonpregnant sheep and changes in the uterine vascular cGMP-dependent pathway and large-conductance Ca²⁺-activated K⁺ channels (BK_Ca). Nonpregnant ewes (n = 19) were synchronized to either follicular or luteal phase using a vaginal progesterone-releasing device (CIDR), followed by intramuscular PGF_2α, CIDR removal, and treatment with pregnant mare serum gonadotropin. UBF was measured with flow probes before tissue collection, and second-generation uterine artery segments were collected from nine follicular and seven luteal phase ewes. The poreforming α- and regulatory β-subunits that constitute the BK_Cα, soluble guanylyl cyclase (sGC), and cGMP-dependent protein kinase G (cPKG) isoforms (cPKG_1α and cPKG_1β) were measured by Western analysis and cGMP levels by RIA. BK_Ca subunits were localized by immunohistochemistry. UBF rose >3-fold (P - < 0.04) in follicular phase ewes, paralleling a 2.3-fold rise in smooth muscle cGMP and 32% increase in cPKG_1α (P < 0.05). sGC, cPKG_1β, and the BK_Ca α-subunit were unchanged. Notably, expression of β₁- and β₂-regulatory subunits rose 51 and 79% (P ≤ 0.05), respectively. Increases in endogenous ovarian estrogens in follicular-phase ewes result in increases in UBF associated with upregulation of the cGMP- and cPKG-dependent pathway and increased vascular BK_Ca β/α-subunit stoichiometry, suggesting enhanced BK_Ca activation contributes to the follicular phase rise in UBF.