Prenatal ethanol exposure decreases GAP-43 phosphorylation and protein kinase C activity in the hippocampus of adult rat offspring.

Consumption of moderate quantities of ethanol during pregnancy produces deficits in long-term potentiation in the hippocampal formation of adult offspring. Protein kinase C (PKC)-mediated phosphorylation of the presynaptic protein GAP-43 is critical for the induction of long-term potentiation. We tested the hypothesis that this system is affected in fetal alcohol-exposed (FAE) rats by measuring GAP-43 phosphorylation and PKC activity in the hippocampus of adult offspring of rat dams that had consumed one of three diets throughout gestation: (a) a 5% ethanol liquid diet, which produced a maternal blood ethanol concentration of 83 mg/dl (FAE); (b) an isocalorically equivalent 0% ethanol diet (pair-fed); or (c) lab chow ad libitum. Western blot analysis using specific antibodies to PKC-phosphorylated GAP-43 revealed that FAE rats had an approximately 50% reduction in the proportion of phosphorylated GAP-43. Similarly, we found that PKC-mediated incorporation of 32P into GAP-43 was reduced by 85% in hippocampal slices from FAE rats compared with both control groups. FAE animals also showed a 50% reduction in total hippocampal PKC activity, whereas the levels of six major PKC isozymes did not change in any of the diet groups. These results suggest that GAP-43 phosphorylation deficits in rats prenatally exposed to moderate levels of ethanol are not due to alterations in the expression of either the enzyme or substrate protein, but rather to a defect in kinase activation.

Influence of maternal grooming, sex and age on Fos immunoreactivity in the preoptic area of neonatal rats: implications for sexual differentiation.

The medial preoptic area (mPOA) of the hypothalamus contains a sexually dimorphic nucleus (SDN-POA) that is 5-7 times larger in males than females and which contributes to the development and expression of male-specific sex behaviors in adulthood. Aside from a critical role for estrogen, the mechanisms that establish and maintain this sex difference are largely unknown. Differences in the size of the SDN-POA are thought to be related to estrogen-associated effects on programmed cell death (apoptosis) during early neonatal development. The expression of male sex behavior is also influenced by maternal behavior during development. During the postnatal period, the dam grooms the anogenital region of the pups to stimulate urination and defecation; however, male pups are groomed significantly more often than females and this maternal attention influences the expression of normal male sexual behavior in adulthood. Based on these observations, we hypothesized that different amounts of anogenital sensory stimulation might contribute to the sexually dimorphic development of the SDN-POA, specifically by providing for different levels of neuronal activation in the SDN-POA resulting in different degrees of cell death. Two experiments were conducted to test this hypothesis. In the first experiment, male and female rat pups on postnatal day 3 (PN 3) received simulated anogenital grooming with a stiff bristle paint brush. One hour later, the brains were removed and sections through the POA were cut and processed for the immunocytochemical detection of Fos-like immunoreactivity (IR) as an indicator of neuronal activation. In the second experiment, male and female littermates were killed on PN 3, 5, 7 and 12 and the number of Fos-immunoreactive cells and pyknotic cells detected in the SDN-POA were counted and compared. Our data demonstrate that anogenital stimulation on PN 3 results in a rapid induction of Fos-immunoreactive in the POA of both males and females. However, the majority of Fos-immunoreactive cells were located in the ventral POA and were distinctly lacking in the SDN-POA. In experiment 2, again no Fos-immunoreactive cells were detected in the SDN-POA of animals examined on PN 5-12. However, there was an increase in the number of pyknotic cells in the area surrounding and including the SDN-POA of females relative to males at PN 5, 7 and 12. Collectively, the data suggest that (1) anogenital grooming during early postnatal development induces a rapid activation of cells in the ventral mPOA, but not in the SDN-POA of rats, (2) there is a greater incidence of cell death in and around the SDN-POA of females vs. males during neonatal development, particularly toward the end of the hormone-sensitive critical period, and (3) Fos expression does not appear to be correlated with the sexually dimorphic development of, and/or programmed cell death within, the developing SDN-POA.