The structure and function of lacrimal and salivary glands present gender differences. Previous works have indicated a synergic action between insulin and androgens over lacrimal gland, and insulin-signaling pathways were recently described in lacrimal gland and salivary gland. Our present study investigates whether gender modulates the early steps of the insulin-signaling system in vivo. Eight-week-old male and female Wistar rats (n = 8/group) were compared to evaluate insulin serum levels and insulin tolerance tests by radioimmunoassay and glucose oxidase method, respectively. To assess insulin receptor (IR), Shc, STAT-1, ERK, and Akt phosphorylation in response to insulin in lacrimal gland and salivary gland, tissues from female and male rats (n = 5-8/group) were submitted to immunoprecipitation and immunoblotting or Western blotting protocol, and phosphorylation level was determined by densitometry. No difference was found in insulin serum levels or insulin tolerance tests comparing both groups. Nevertheless, lacrimal gland and salivary gland of female rats had a significantly lower insulin-induced IR phosphorylation compared with males. IR phosphorylation was not affected by the estrous cycle stage in either tissue. In addition, in females an apparent but not significant lower STAT and Akt phosphorylation in response to insulin was observed in the lacrimal gland, compared with males. Our findings suggest that alterations in insulin signal transduction may play a role in lacrimal gland and salivary gland gender differences.