Fig 1.
Representative images of vaginal smears from mice in estrus and diestrus.
(A) Image showing a predominance of leukocytes in diestrus. (B) Image showing a predominance of anucleated epithelial cells in estrus.
Fig 2.
Increased sensitivity of VTA DA neurons to inhibition by DA during diestrus.
Extracellular recordings of VTA DA neurons were obtained from mice in diestrus and estrus. The DA response was measured in putative VTA DA neurons with 0.5, 1, 2, 5, and 10 μM DA. (A-B) Representative rate meter graphs show the effects of DA during (A) diestrus and (B) estrus. (C) Concentration-response graph showing enhanced inhibition to DA in mice during diestrus (n = 9) compared with estrus (n = 9). *P < 0.05 by two-way ANOVA.
Fig 3.
Increased sensitivity of VTA DA neurons to ethanol excitation during diestrus.
Extracellular recordings of VTA DA neurons were obtained from mice in diestrus and estrus. The ethanol response was measured in VTA DA neurons with 40, 80, and 120 mM ethanol. Representative rate meter graphs show the effects of ethanol during (A) diestrus and (B) estrus. (C) Pooled concentration-response graph showing excitation by ethanol in mice during diestrus (n = 9) compared with estrus (n = 7). **P < 0.01 by two-way ANOVA.
Fig 4.
E2 enhances DA inhibition and ethanol excitation of VTA DA neurons.
Extracellular recordings were obtained from VTA DA neurons in ovariectomized (OVX) mice previously treated with 17β-estradiol-3-benzoate (EB) or vehicle (VEH) (A-B) Representative rate meter graphs showing response to 0.5–10 μM DA of a VTA DA neuron from a mouse treated with VEH (A) or EB (B). (C) DA responses in VTA DA neurons treated with 0.5–10 μM DA. Concentration-response graph shows enhanced inhibition by DA in OVX mice treated with EB compared with VEH (n = 9 per group). (D-E) Representative rate meter graphs showing response to 40–120 mM ethanol of a VTA DA neuron from a mouse treated with VEH (D) or EB (E). (F) Ethanol responses in VTA DA neurons treated with 40, 80, and 120 mM ethanol. Pooled concentration-response graph shows enhanced excitation to ethanol in OVX mice treated with EB (n = 13) compared with VEH (n = 12). *P < 0.05, **P < 0.01 by two-way ANOVA.
Fig 5.
Estrogen receptors in the VTA acutely regulate the sensitivity of DA neurons to ethanol but not DA.
(A-B) Representative rate meter graphs showing response to 5 μM DA of a VTA DA neuron from a mouse in estrus (A) or diestrus (B) recorded before and during treatment with ICI 182,780. (C) Response to 5 μM DA of VTA DA neurons from mice in estrus (n = 6) or diestrus (n = 6) before and after treatment of slices containing the VTA with ICI 182,780. There was no change in DA inhibition with ICI 182,780 treatment, but there was a significant effect of cycle phase (*P < 0.05). (D-E) Representative rate meter graphs showing response to 80 mM ethanol of a VTA DA neuron from a mouse in estrus (D) or diestrus (E) recorded before and during treatment with ICI 182,780. (F) Response to 80 mM ethanol of VTA DA neurons from mice in estrus (n = 6) or diestrus (n = 6) before and after treatment of slices containing the VTA with ICI 182,780. The enhanced VTA DA neuron excitation by ethanol that was observed in mice in diestrus was decreased after treatment with ICI 182,780, whereas the ethanol response of VTA DA neurons from mice in estrus was not altered by ICI 182,780. *P < 0.05 by two-way RM ANOVA. #P < 0.05 between mice in estrus and diestrus before ICI 182,780 treatment by post-hoc Sidak’s multiple comparisons test.