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(Investigative Ophthalmology and Visual Science. 2000;41:1142-1148.)
© 2000 by The Association for Research in Vision and Ophthalmology, Inc.

Cholinergic and Adrenergic Modulation of the Ca2+ Response to Endothelin-1 in Human Ciliary Muscle Cells

Ganesh Prasanna1, Adnan I. Dibas2 and Thomas Yorio1

1 From the Department of Pharmacology, University of North Texas Health Science Center, Fort Worth, Texas; and the 2 Department of Immunology, St. Paul Medical Center, Mary Kay Ash Institute for Cancer Research, Dallas, Texas.

PURPOSE. To determine the cholinergic (carbachol, CCH) and adrenergic (norepinephrine, NE) modulation of Ca2+ response to endothelin-1 in human ciliary smooth muscle (HCSM) cells.

METHODS. Intracellular calcium levels were measured using the Fura-2 calcium imaging system in HCSM cells treated either singly with endothelin-1 (ET-1; 2–200 nM), CCH (1–100 µM), NE (0.1–10 µM) or isoproterenol (ISO; 1 µM) or in combinations of CCH, NE, or ISO with ET-1. Intracellular cAMP levels after NE and ISO treatments were also measured using a radioimmunoassay.

RESULTS. Endothelin-1 dose-dependently increased [Ca2+]i and was characteristically biphasic (peak [Ca2+]i for ET-1: 2 nM, 517 ± 73 nM; 20 nM, 785 ± 65 nM; and 200 nM, 2564 ± 359 nM). Carbachol also dose-dependently increased [Ca2+]i; however, subsequent additions of ET-1 (200 nM) resulted in lower [Ca2+]i (100 µM CCH + ET-1; 300 ± 21 nM) compared with that observed with 200 nM ET-1 alone (2564 ± 359 nM). Norepinephrine pretreatment also decreased ET-1–induced [Ca2+]i (10 µM NE + ET-1; 619 ± 64 nM) compared with ET-1 alone, and NE’s effect could be reversed by propranolol (ß-adrenergic antagonist) treatment. Neither CCH nor NE was able to completely abolish ET-1’s ability to mobilize calcium in HCSM cells. Isoproterenol (a ß-agonist) mimicked NE’s effect on ET-1–induced [Ca2+]i (1 µM ISO + ET-1; 254 ± 56 nM). Both ISO and NE elevated [cAMP] in HCSM cells.

CONCLUSIONS. In HCSM cells, CCH and ET-1 can activate common as well as specific [Ca2+]i pools. The reduction in ET-1–induced [Ca2+]i after NE/ISO treatment appears to be due to elevated cAMP levels via ß-receptor activation, suggesting the existence of receptor cross talk. The ability of CCH and NE to modulate ET-1’s actions on HCSM may be relevant to the regulation of ciliary muscle contraction and aqueous humor outflow.




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