Fifty reproductively mature female pinealectomized or sham-operated white-footed mice (Peromyscus leucopus) were injected daily for 7 weeks with 50 micrograms melatonin at 12 hr after lights on. The injections started either on the day following surgery or 2 weeks after surgery. All mice were maintained at room temperature under long-day photoperiod (LD 16:8). Pinealectomized and sham-operated mice injected immediately following surgery and sham-operated mice injected beginning 2 weeks after surgery had reduced reproductive tract weights (30 +/- 6, 33 +/- 6, and 33 +/- 3 mg, respectively) compared to the pinealectomized mice injected beginning 2 weeks after surgery (78 +/- 8 mg). In a second study 74 pinealectomized or sham-operated mice were injected at either 2, 7.5, 12, or 15 hr after lights on with 50 micrograms melatonin. Irrespective of surgical treatment, mice injected at 15 hr had a mean (+/- SE) reproductive tract weight (33 +/- 10 mg) which was substantially less than the 2-hr group (78 +/- 17 mg) while the 7.5- and 12-hr groups were intermediate (P less than 0.001). Pinealectomized and sham-operated animals responded similarly to melatonin injections at these four time points (P less than 0.35). The clearance rate of plasma melatonin after a single 50-micrograms melatonin injection or after 2 weeks of daily injections at either 2 or 12 hr after lights on was determined. Melatonin concentrations were determined using high-performance liquid chromatography and radioimmunoassay.(ABSTRACT TRUNCATED AT 250 WORDS)