Alpha-2 Adrenoceptor Mediated Changes in Aqueous Dynamics: Effect of Pertussis Toxin

Abstract

Because alpha-2 (α2) adrenoceptor-mediated inhibition of signal transduction mechanisms is regulated, in part, by inhibitory G (G1) protein, we studied the effects of pertussis toxin (PTX) pretreatment on α2 adrenergic agonist, UK 14.304-18 (UK; brimonidine)-induced: (1) changes in intraocular pressure (IOP) and aqueous flow rate of rabbits; (2) modulation of 3H-norepinephrine (NE) overflow of rabbit iris-ciliary bodies (ICBs) and (3) accumulation of cyclic AMP in rabbit ICBs and cultured non-pigmented ciliary epithelial (NPE) cells. Results showed that UK (50 μg) lowered the IOP of normal rabbits by 8 ± 1 mmHg (n = 8) at 3 hr when treated topically and that the reduction of IOP was accompanied by a decrease in aqueous humor inflow (35%, n = 5). Therefore, it was postulated that these UK-induced effects involve activation of a G1 protein linked to α2 adrenoceptors. In PTX-pretreated (2·5 μg kg-1, i.a.) rabbits, hypotensive responses to UK were reduced by 50% and 70% (n = 8) at days 4 and 7 post PTX treatment, respectively. The suppression of aqueous humor inflow induced by UK was also prevented by the PTX treatment. In isolated, perfused rabbit ICBs, UK (1 μM) caused 50% inhibition of 3 H-NE overflow from electrical field stimulation. Pretreatment with PTX (150 ng ml-1, 4 hr) partially prevented UK-induced inhibition of NE overflow. In in vitro assays of postjunctional α2 adrenoceptor activity, UK (l μM) inhibited isoproterenol (ISO, 1 μM)-stimulated cAMP accumulation by 45% and 48% in ICBs and NPE cells, respectively. Furthermore, pretreatment with PTX (150 ng ml-1, 4 hr on ICBs and 100 ng ml--1, 18 hr on NPE cells) prevented UK-induced inhibition of ISO-stimulated cAMP accumulation in NPE cells. These in vivo and in vitro data confirm that G1 protein(s) are involved in the regulation of aqueous humor dynamics by α2 adrenoceptors.