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The rodent cerebellum is richly supplied with PACAPergic innervation. Exogenous pituitary adenylate cyclase-activating polypeptide (PACAP) increases cerebellar granule cell survival and differentiation in culture, and enhances the number of neuroblasts in the molecular and internal granule cell layers (IGL) when injected postnatally into the cerebellum in vivo. Here, we have investigated the role of endogenous PACAP during cerebellar development by comparing the morphology of normal and PACAP-deficient mouse cerebellum, and the response of cerebellar granule cells from normal and PACAP-deficient mice subjected to neurotoxic insult in culture. There was no difference in cerebellar volume or granule cell number, in 11-day-old wild type versus PACAP-deficient mice. Cultured cerebellar neurons from PACAP-deficient and wild type mice also showed no apparent differences in survival and differentiation either under depolarizing conditions, or non-depolarizing conditions in the presence or absence of either dibutyryl cAMP or 100 nM PACAP. However, cultured cerebellar neurons from PACAP-deficient mice were significantly more sensitive than wild type neurons to ethanol- or hydrogen peroxide-induced toxicity. Differential ethanol toxicity was reversed by addition of 100 nM exogenous PACAP, suggesting that endogenous PACAP has neuroprotective activity in the context of cellular insult or stress. The neuroprotective action of PACAP was mimicked by dibutryl cAMP, indicating that it occurred via activation of adenylate cyclase. These results indicate that PACAP might act to protect the brain from paraphysiological insult, including exposure to toxins or hypoxia.

Jegou et al. (2012) have reported prenatal alcohol exposure (PAE)-induced reductions of angiogenesis-related proteins in mouse placenta. These effects were associated with striking alterations in microvascular development in neonatal cerebral cortex. Here, we employed a rat model of moderate PAE to search for additional proteins whose placental and fetal cortical expression is altered by PAE, along with a subsequent examination of fetal cerebral cortical alterations associated with altered protein expression. Long-Evans rat dams voluntarily consumed either a 0 or 5% ethanol solution 4 h each day throughout gestation. Daily ethanol consumption, which resulted in a mean peak maternal serum ethanol concentration of 60.8 mg/dL, did not affect maternal weight gain, litter size, or placental or fetal body weight. On gestational day 20, rat placental: fetal units were removed by Caesarian section. Placental protein expression, analyzed by 2D-PAGE - tandem mass spectroscopy, identified a total of 1,117 protein spots, 20 of which were significantly altered by PAE. To date, 14 of these PAE-altered proteins have been identified. Western blotting confirmed the alterations of two of these placental proteins, namely, annexin-A4 (ANX-A4) and cerebral cavernous malformation protein 3 (CCM-3). Specifically, PAE elevated ANX-A4 and decreased CCM-3 in placenta. Subsequently, these two proteins were measured in fetal cerebral cortex, along with radiohistochemical studies of VEGF binding and histofluorescence studies of microvascular density in fetal cerebral cortex. PAE elevated ANX-A4 and decreased CCM-3 in fetal cerebral cortex, in a pattern similar to the alterations observed in placenta. Further, both VEGF receptor binding and microvascular density and orientation, measures that are sensitive to reduced CCM-3 expression in developing brain, were significantly reduced in the ventricular zone of fetal cerebral cortex. These results suggest that the expression angiogenesis-related proteins in placenta might serve as a biomarker of ethanol-induced alterations in microvascular development in fetal brain.

AbstractTransient exposure to ethanol (EtOH) results in a massive neurodegeneration in the developing brain leading to behavioral and cognitive deficits observed in fetal alcohol syndrome. There is now compelling evidence that K+ channels play an important role in the control of programmed cell death. The aim of the present work was to investigate the involvement of K+ channels in the EtOH‐induced cerebellar granule cell death and/or survival. At low and high concentrations, EtOH evoked membrane depolarization and hyperpolarization, respectively. Bath perfusion of EtOH (10 mM) depressed the IA (transient K+ current) potassium current whereas EtOH (400 mM) provoked a marked potentiation of the specific IK (delayed rectifier K+ current) current. Pipette dialysis with GTPγS or GDPβS did not modify the effects of EtOH (400 mM) on both membrane potential and IK current. In contrast, the reversible depolarization and slowly recovering inhibition of IA induced by EtOH (10 mM) became irreversible in the presence of GTPγS. EtOH (400 mM) induced prodeath responses whereas EtOH (10 mM) and K+ channel blockers promoted cell survival. Altogether, these results indicate that in cerebellar granule cells, EtOH mediates a dual effect on K+ currents partly involved in the control of granule cell death.