Six6^flox/flox/LHRH^cre females are hypogonadotropic because of deficient GnRH. (a) Serum was collected from adult Six6^flox/flox (Cre−) and Six6^flox/flox/LHRH^cre (Cre+) female diestrus mice and assayed for (a) LH (n = 5 or 6) and (b) FSH (n = 5 or 6). (a and b) Statistical analysis by Student t test as compared with Cre−. **P < 0.01. (c) Serum LH was measured immediately before and 10 min after a single GnRH challenge in Cre− and Cre+ diestrus females (n = 5 or 6). Numerical values on the graphs indicate the average fold-change of Cre− and Cre+ mice LH in response to GnRH injections. Statistical analysis was by two-way repeated-measures ANOVA [there was a statistically significant effect of kisspeptin injection on LH: F(1, 7) = 51.82, P = 0.0002]. (d) Serum LH was measured before and 15 minutes after a single kisspeptin challenge in Cre− and Cre+ diestrus females (n = 5 or 6). Numerical values on the graphs indicate the average fold-change of Cre− and Cre+ mice LH in response to kisspeptin injections. Statistical analysis was by two-way repeated-measures ANOVA. [There was a statistically significant effect of kisspeptin injection on LH, F(1, 6) = 11.09, P = 0.016; there was a statistically significant effect of genotype on LH, F(1, 6) = 48.62, P = 0.0004.]