This study sought to examine gender differences in emotional reactions and compliance with Ministry of Health (MOH) guidelines during the COVID-19 pandemic in Israel, with the goal of gaining a deeper understanding of these gender-related variations throughout the lockdown periods. A longitudinal study comprising 2509 participants was conducted during two of Israel's lockdowns: 1424 participants completed a questionnaire during the first lockdown (23 April-5 May 2020); of these, 1085 completed a follow-up questionnaire during the second lockdown (September 30-October 10, 2020). Participants exhibited higher levels of compliance with MOH guidelines (e.g., wearing face masks, maintaining social distancing) and knowledge about COVID-19 during the second lockdown, whereas they exhibited more negative emotional reactions during the first lockdown. Female participants scored higher than male participants on all measures. Multiple regression results showed that about 21% of the variance in compliance with MOH guidelines was explained by lockdown type (i.e., first or second), gender, and age, while knowledge and negative emotional reactions added another 19% to the explained variance. The results suggest that the impact of the pandemic on emotional reactions decreased over time, with people exhibiting greater compliance with MOH guidelines and more knowledge about COVID-19. Moreover, the behavioral and psychological impact of the pandemic was greater on women than on men. The results suggest that healthcare professionals should pay more attention to mental health issues during a pandemic. Moreover, policymakers should focus on women as a vulnerable group and suggest appropriate solutions to reduce their emotional distress. Furthermore, governments and employers should provide greater flexibility and support for single mothers during the pandemic. In addition, gender inequality during lockdowns may place women at greater risk of psychological distress.