An in vitro perifusion system was used to investigate the spontaneous luteinizing hormone (LH) release from 10 human fetal (21–23 weeks of gestation) and 1 adult female pituitaries. The pattern of LH release from fetal pituitaries (n = 6) exhibited a remarkable pulsatile character with a mean (±SE) pulse interval of 12.7 + 1.7 min. The mean pulse amplitude was 5.2 ± 0.9 mlU with a nadir to peak increment of 69.5 ± 6.4%. The mean LH release rate was 12.3 ± 3.3 mIU/2 min. Blockade of calcium activity with 0.1 mM verapamil and 4 mM EGTA suppressed the frequency (from 1 pulse/12–20 min to 1 pulse/50–100 min) and amplitude (from 5.4–5.7 mlU to 1.4–2.1 mlU) of this spontaneous pulsatile LH release (n = 2). Administration of 8 nMgonadotropin-releasing hormone induced 255 and 954% increases in LH secretion (n = 2). Each quarter of an adult human pituitary also secreted LH in a pulsatile fashion, with a pulse interval of 15.2 ± 5.6 min, a pulse amplitude of 5.4 ± 0.6 mlU, a nadir to peak increment of 67.5 ± 5.2%, and an overall release rate of 14.8 ± 0.9 mIU/2 min. These studies demonstrate that LH release from the isolated human pituitary in vitro is characterized by high-frequency/low-amplitude pulses, independent of hypothalamic stimulation. Accordingly, this spontaneous calcium-mediated pulsatile LH release apparently reflects the activity of an intrinsic intrapituitary pulse-generating mechanism.