Changes in cytosolic free calcium concentration ([Ca2+]i) in response to cholinergic and adrenergic agents alone and in combination were investigated using fura-2 fluorescence imaging in intact non-pigmented epithelial cells of rabbit ciliary body. 2. Resting ('baseline') [Ca2+]i was 147 +/- 6 nM (mean +/- S.E.M.). Acetylcholine (ACh, 10 microM) doubled [Ca2+]i, and adrenaline (1 microM) increased it by about 36%. When ACh (10 microM) and adrenaline (1 microM) were applied together [Ca2+]i was transiently increased to 1160 +/- 160 nM, about 7 times the response induced by ACh alone. 3. Noradrenaline and 5-bromo-6-(2-imidazolin-2-yl-amino)-quinoxaline (UK 14304) had effects similar to adrenaline in enhancing the response to ACh. Phenylephrine (Phe) had a relatively smaller effect and none was observed for methoxamine and isoprenaline (Iso). 4. The response to ACh and adrenaline could be blocked by atropine (1 microM, 87 +/- 5%), yohimbine (1 microM, 73 +/- 8%), and to a lesser degree by prazosin (1 microM). Propranolol had no effect. 5. Lowering the extracellular calcium concentration to 3 nM dropped the baseline [Ca2+]i by half and reduced the response to ACh and adrenaline to a small and transient rise in [Ca2+]i. Addition of La3+ to Ca(2+)-containing solution also lowered [Ca2+]i and largely reduced the response. 6. We conclude that simultaneous activation of muscarinic and alpha 2-adrenergic receptors induces a large increase in [Ca2+]i, which is the result of both Ca2+ release and influx.
