The effects of prostaglandins E2 and F2α on porcine ciliary muscle cells in culture

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Abstract
The purpose of this work is to investigate the effect of prostaglandins (PGs) on the contraction of ciliary muscle cells. It has been proposed that PGs induce relaxation of ciliary muscle and facilitate uveoscleral outflow, and reduce intraocular pressure (IOP). The ocular response to PGs is complicated because the relative contributions of uveoscleral flow and the conventional outflow to lowering IOP and the type of PG receptors associated with ciliary muscle may vary depending on animal species. In order to obtain insights into prostaglandin receptors of ciliary muscle, ciliary muscle cells from porcine eye were grown in culture and characterized immunocytochemically with antibodies against smooth muscle-alpha-actin and PGE2 receptor subtypes. As in ciliary muscle tissues, positive immunostaining for alpha-actin and EP2 and EP3 subtypes was observed in cultured cells. Time-dependent contraction of cultured cells induced by 10(-4) M carbachol was recorded by taking sequential photographs and analyzed. Using this assay method, the effect of prostaglandins E2 and F2 alpha to inhibit the carbachol-induced contraction was studied. PGE2 showed potent inhibition of cell contraction; 10(-7) approximately 10(-8) M PGE2 inhibited 50% of full contraction in 15 min. PGF2 alpha at 10(-4) M neither caused cell contraction by itself nor blocked carbachol-induced contraction. The EP2 agonist 11-deoxy-16, 16-dimethyl PGE2 at 10(-4) M inhibited cell contraction but the EP3 agonist sulprostone had no effect. Dibutyryl cAMP at 3 x 10(-5) M inhibited contraction by 50%. In the presence of the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (IBMX), less than 10(-7) M dibutyryl cAMP caused 50% inhibition. In support of the cAMP effect, the addition of 10(-4) M PGE2 to cultured cells in the presence of indomethacin and IBMX was shown to cause an 80% increase in intracellular cAMP concentration compared with the basal (i.e. unstimulated) level of cAMP. Stimulation of cells with 10(-4) M PGF2 alpha caused no increase in cellular cAMP. These results indicate that PGE2 receptor EP2 subtype, but not PGF2 alpha receptor, is involved in the inhibition (hence relaxation by inference) of carbachol-induced porcine ciliary muscle cell contraction. It awaits further studies to determine whether cultured ciliary muscle cells of other species respond similarly to different PGs.