Intracellular Ca²⁺ transients occur during neurotransmitter release from postganglionic autonomic nerve terminals, and during smooth muscle cell contraction, yet in both of these cases the pathways for generating these transients, and their relationship with function, are not clear. The anococcygeus muscle is a smooth muscle preparation that is currently little studied, which offers the opportunity to study noradrenergic and nitrergic neurotransmission, which in this tissue control contraction and relaxation respectively. Experiments were performed on anococcygeus muscles isolated from male Wistar rats (200-400 g). Rats were killed humanely. The anococcygeus was excised and incubated in the cell-permable acetylmethoxy ester (AM) form of the Ca²⁺ indicator Oregon Green BAPTA-1 for 70 min at 35°C, before being transferred to a superfused stage on a laser scanning confocal microscope (Leica DM-LFSA SP2). The indicator accumulated in smooth muscle cells and in structures resembling nerve bundles. Drugs were applied by addition to the perfusion reservoir. Electrical field stimulation (1-10 pulses, 1-10 Hz, pulse duration 0.05-0.2 ms, at supramaximal voltage) evoked Ca²⁺ transients in the nerve bundles, which preceded contraction. Trains of electrical pulses also elicited three distinct types of Ca²⁺ transients in smooth muscle cells: fast (usually 1000 ms); and propagating waves, which spread (at a mean speed of 49 ± 5 (S.E.M.) μm s^-1; n = 6 cells) along the length of the smooth muscle cells. Prazosin (100 nM), an α₁-adrenoceptor antagonist, did not affect electrically-evoked Ca²⁺ transients in the nerve bundles nor the fast localised transients but abolished waves and whole cell Ca²⁺ transients in the smooth muscle cells, and eliminated contraction (n = 9 preparations). Tetrodotoxin (1 μM), a voltage-gated Na⁺ channel blocker, abolished electrically evoked Ca²⁺ transients in both nerve bundles and smooth muscle cells, and eliminated contraction (n = 5 preparations). α,β-methylene ATP (1 μM), a P2X receptor agonist, elicted localised Ca²⁺ transients in smooth muscle cells, and following continued exposure abolished the fast, localised transients produced during electrical stimulation (n = 3 preparations). These results indicate that neurotransmitter release in the anococcygeus elicits fast, focal purinergic Ca²⁺ transients, as well as Ca²⁺ waves and whole cell transients initiated by noradrenaline.