Information on the relative activity and on the functional relationships between the acutely releasable (1st) and reserve (2nd) pools of pituitary LH during the course of the normal menstrual cycle was obtained via a 4 h LRF infusion (0.2μg/min × 4 h). This was immediately followed by 3 pulses of LRF (10 μg at 2 h intervals) to assess further the size of the acutely releasable pool after activation of the reserve pool by the infusion. Our observations indicate that two functional pools of LH are present in all phases of the menstrual cycle and that comparative pool size or activity is influenced profoundly by ovarian steroid feedback as well as by the pattern of input of hypothalamic LRF. From the early to the late follicular phase, in synchrony with the rising levels of E2, the size of the 2nd pool is preferentially augmented. A small increase in the 1st pool activity is not apparent until the late follicular phase when a 5-fold increase in the size of the 2nd pool is also attained. During the mid-luteal phase and in association with relatively high progesterone (P) and E2, the large 2nd pool is maintained as in the late follicular phase but the 1st pool is strikingly smaller. Activation of the 2nd pool of LH by the LRF infusion (priming) increases the acutely releasable LH (1st pool) in all three phases of the cycle, as evidenced by an enhanced response to the 1st but not subsequent pulses of LRF at the end of the infusion as compared with non-infused controls. This priming effect is likely a reflection of activation or “shifting” of LH from the larger 2nd pool to the smaller 1st pool. It is found that this priming effect is greatest during the mid-luteal phase as compared to other phases of the cycle. During the days of mid-cycle LH surge, a dramatic reversal of the relative activity of the two pools is observed and this is manifested by an enormous increase in the activity of the 1st relative to the 2nd pool. In contrast to other phases of the cycle, the release of LH from the 2nd pool is not sustained and this premature decline in LH release despite continuous LRF infusion appears to be due to pituitary depletion of LH as evidenced by the failure of the pituitary response to pulses of LRF immediately following the infusion. These findings together with our previous observations suggest that 1) LRF not only induces synthesis- storage (2nd pool) and release (1st pool) of LH, but also activates the 2nd pool and renders its LH more readily releasable; 2) these positive influences exerted by LRF are amplified by the presence of E2 which appears to provide a “permissive action” of LRF except E2 also functions to impede the LRF mediated LH-release; 3) the mid-luteal levels of P do not inhibit the augmented 2nd pool due to E2 but may in fact amplify the LRF induced activation of the 2nd pool with the enlargement of the smaller 1st pool; and 4) at mid-cycle, the assumed increased endogenous LRF release together with the development of estrogen dependent self-priming effect of LRF induces a dramatic shifting of LH from the 2nd to the 1st pool with accelerated LH release by overcoming the impeding action of E2.