Interactions between forskolin, GS, and divalent cations on ciliary process adenylyl cyclase and intraocular pressure in the rabbit eye.

The major adenylyl cyclase activity (AC) of rabbit ocular ciliary processes was investigated by dose-response analysis with respect to interactions of its regulators [the stimulatory G-protein (alpha S) and forskolin] under both saturating and suboptimal divalent cation (M2+) conditions. alpha S was generated directly with fluoroaluminate or via receptors for vasoactive intestinal peptide or isoproterenol. Forskolin and alpha S when liganded simultaneously to AC at suboptimal M2+, potentiated maximal enzyme activity (approximately twice the sum of the separate activities), while the responses were additive (no potentiation) under saturating M2+ conditions. The results demonstrate that the three ligands (forskolin, M2+ and alpha S) mutually interact in a cooperative manner as follows: (1) The apparent k(act) of forskolin for AC was decreased by 1.7-1.9 log units when alpha S was liganded to AC. (2) Conversely, with forskolin liganded to AC the apparent k(act) of alpha S for AC also decreased by a factor of at least 3-4. (3) The nature of the divalent cation M2+ had no effect on the k(act) of alpha S for AC alone and for the AC:FSK complex, and also did not affect the apparent k(act) of forskolin for AC alone, or for the AC: alpha S complex. The nature of M2+ (Mg2+ or Mn2+) affected only the maximal catalytic rate (Emax). (4) Potentiation of adenylyl cyclase by alpha S + FSK was demonstrable in vivo by the increased ocular pressure response of the rabbit eye when a low topical dose of isoproterenol (125 ng per eye) and a subthreshold dose of forskolin (100 micrograms per eye) were given in combination.