Binge alcohol-induced alterations in BDNF and GDNF expression in central extended amygdala and pyriform cortex on infant rats.

Mothers who consume alcohol during pregnancy may cause a neurotoxic syndrome termed fetal alcohol spectrum disorder (FASD) in the offspring, which includes cognitive deficits and emotional/social disturbances. These alterations are thought to be caused, at least in part, by alcohol-induced imbalance in neurotrophic factor levels, which are critically involved in normal neurodevelopment. Our goal was to study whether brain-derived neurotrophic factor (BDNF) and glial-derived neurotrophic factor (GDNF) expression were affected by alcohol in central extended amygdala (CEXA) and pyriform cortex (Pyr), structures strongly involved in emotional/social behaviors. Further, we evaluated how these changes could be related to blood and brain alcohol concentrations. Postnatal day (PND) pups at 7, 15 and 20-days old were administered alcohol (2.5g/kg s.c. at 0 and 2h) or saline. Immunohistochemistry was used to detect the expression of BDNF and GDNF at 2, 12 and 24h after drug administration. Also, gas chromatography was bused to measure blood alcohol levels (BALs) and brain alcohol levels (BrALs) at each hour, from 2 to 8h after the second alcohol administration. Results showed: (1) alcohol-induced enhancement of BDNF positive cells on PND 7 and 20, a decrease on PND 15 in the CEXA, and no changes in the Pyr on PND 7 and 20, but a diminished on PND 15; (2) GDNF positive cells rise after alcohol administration for the three ages in the CEXA and Pyr except on PND 15, where there was a decline; and (3) pharmacokinetics analysis demonstrated age-related differences showing equal BALs on PND 7 and 20 but higher BALs on PND 15. In contrast, BrALs were higher on PND 7 than 15 and 20. Hence, BALs may not be predictive of BrALs in postnatal rats. Furthermore, we did not find a relationship between age in pharmacokinetic differences and neurotrophins response. In conclusion, the CEXA and Pyr are brain structures sensitive to alcohol-induced imbalance in neurotrophic factors expression; and BALs are not a mirror of BrALs.

Detection of conspecific pheromones elicits fos expression in GABA and calcium-binding cells of the rat vomeronasal system–medial extended amygdala

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The olfactory accessory system is specialized in the detection of pheromones, being an afferent to medial extended amygdala. In spite of the fact that numerous phenotypes are found in these structures, in the current literature, there are no detailed descriptions about the phenotype of neurons in the vomeronasal system–medial extended amygdale after their activation by pheromonal stimuli. Using immunohistochemistry for fos and dual immunohistochemistry for fos and phenotypes, here we show that females have a greater number of activated neurons by the pheromonal stimulus. Likewise, a great colocalization of fos with GABA, calretinin, and calbindin was observed in the vomeronasal system–medial extended amygdala. These data suggest that in amygdaloid areas, neuronal excitability is controlled by GABAergic neurons that contain different calcium-binding proteins, indicating the important role of inhibitory control on the incoming sensory pheromonal and olfactory inputs controlled and processed by the vomeronasal system (AU)

Kainic acid-induced early genes activation and neuronal death in the medial extended amygdala of rats.

The medial extended amygdala modulates pheromonal perception, influencing emotional and social behavior. As the amygdala is part of neuronal circuits that are very sensitive to excitability, its neurons are targets of seizures in temporal lobe epilepsy. It has been suggested that the hippocampus is strongly involved this pathology. There is less consistent information, however, on the effects of this disease in the amygdala. The effects of status epilepticus on the medial extended amygdala were analyzed by immunohistochemistry for neural stress and by the amino-cupric-silver technique for neuronal death in rats after kainic acid (KA) administration. Sixty adult Wistar male rats were used. Thirty animals received an injection of KA, and 30 were injected with saline. After 2, 4, 12, 24 and 48 h survival the brains were stained for Fos and FosB and for neuronal death. In the present study we show that KA induces Fos and FosB expression in neurons of the medial extended amygdala after 2, 4-48 h, with time courses that are different between them and from control animals. While Fos-IR peaks at 2-4 h post KA and then decreases, FosB-IR increases in the same period reaching its highest expression at 24-48 h. Moreover, KA injection produced massive neuronal death with a peak at 24 h. This neurodegeneration paralleled FosB-IR protein expression. These findings show that KA produces neuronal stress and activation of early genes and neuronal death in the medial extended amygdala, demonstrating the vulnerability of its neurons to the epileptogenic effects of KA.

Detection of conspecific pheromones elicits fos expression in GABA and calcium-binding cells of the rat vomeronasal system-medial extended amygdala.

The olfactory accessory system is specialized in the detection of pheromones, being an afferent to medial extended amygdala. In spite of the fact that numerous phenotypes are found in these structures, in the current literature, there are no detailed descriptions about the phenotype of neurons in the vomeronasal system-medial extended amygdala after their activation by pheromonal stimuli. Using immunohistochemistry for fos and dual immunohistochemistry for fos and phenotypes, here we show that females have a greater number of activated neurons by the pheromonal stimulus. Likewise, a great colocalization of fos with GABA, calretinin, and calbindin was observed in the vomeronasal system-medial extended amygdala. These data suggest that in amygdaloid areas, neuronal excitability is controlled by GABAergic neurons that contain different calcium-binding proteins, indicating the important role of inhibitory control on the incoming sensory pheromonal and olfactory inputs controlled and processed by the vomeronasal system.

Timed changes of synaptic zinc, synaptophysin and MAP2 in medial extended amygdala of epileptic animals are suggestive of reactive neuroplasticity.

Repeated seizures induce permanent alterations of the brain in experimental models and patients with intractable temporal lobe epilepsy (TLE), which is a common form of epilepsy in humans. Together with cell loss and gliosis in many brain regions, synaptic reorganization is observed principally in the hippocampus. However, in the amygdala this synaptic reorganization has been not studied. The changes in Zn density, synaptophysin and MAP(2) as markers of reactive synaptogenesis in medial extended amygdala induced by kainic acid (KA) as a model of TLE was studied. Adult male rats (n=6) were perfused at 10 days, 1, 2, 3 and 4 months after KA i.p. injection (9 mg/kg). Controls were injected with saline. The brains were processed by the Timm's method to reveal synaptic Zn and analyzed by densitometry. Immunohistochemistry was used to reveal synaptophysin and MAP(2) expression. A two-way ANOVA was used for statistics, with a P<0.05 as a significance limit. Normal dark staining was seen in all medial extended amygdala subdivisions of control animals. At 10 days post KA injection a dramatic loss of staining was observed. A slow but steady recovery of Zn density can be followed in the 4 month period studied. Parallel, from 10 days to 2 months stronger synaptophysin expression could be observed, whereas MAP(2) expression increased from 1 month with peak levels at 3-4 months. The results suggest that a process of sprouting exists in surviving neurons of medial extended amygdala after status epilepticus and that these neurons might be an evidence of a reactive synaptogenesis process.