Pierluigi Olla

Chronic vagus nerve stimulation induces neuronal plasticity in the rat hippocampus

Vagus nerve stimulation (VNS) is used to treat pharmacotherapy-resistant epilepsy and depression. However, the mechanisms underlying the therapeutic efficacy of VNS remain unclear. We examined the effects of VNS on hippocampal neuronal plasticity and behaviour in rats. Cell proliferation in the hippocampus of rats subjected to acute (3 h) or chronic (1 month) VNS was examined by injection of bromodeoxyuridine (BrdU) and immunohistochemistry. Expression of doublecortin (DCX) and brain-derived neurotrophic factor (BDNF) was evaluated by immunofluorescence staining. The dendritic morphology of DCX+ neurons was measured by Sholl analysis. Our results show that acute VNS induced an increase in the number of BrdU+ cells in the dentate gyrus that was apparent 24 h and 3 wk after treatment. It also induced long-lasting increases in the amount of DCX immunoreactivity and in the number of DCX+ neurons. Neither the number of BrdU+ cells nor the amount of DCX immunoreactivity was increased 3 wk after the cessation of chronic VNS. Chronic VNS induced long-lasting increases in the amount of BDNF immunoreactivity and the number of BDNF+ cells as well as in the dendritic complexity of DCX+ neurons in the hippocampus. In contrast to chronic imipramine treatment, chronic VNS had no effect on the behaviour of rats in the forced swim or elevated plus-maze tests. Both chronic and acute VNS induced persistent changes in hippocampal neurons that may play a key role in the therapeutic efficacy of VNS. However, these changes were not associated with evident behavioural alterations characteristic of an antidepressant or anxiolytic action.

Voluntary ethanol consumption induced by social isolation reverses the increase of α 4 /δ GABA A receptor gene expression and function in the hippocampus of C57BL/6J mice

Post-weaning social isolation (SI) is a model of prolonged mild stress characterized by behavioral and neurochemical alterations. We used SI in C57BL/6J mice to investigate the effects of ethanol (EtOH) in the free-choice drinking paradigm on gene expression and function of γ-aminobutyric acid type A receptors (GABAARs) and the role of neuroactive steroids in the actions of EtOH in the hippocampus. SI stress induced a marked reduction in hippocampal 3α-hydroxy-5α-pregnan-20-one (3α,5α-TH PROG) and was associated with molecular and functional changes of the GABAAR. The gene expression of the α4 and δ subunits was increased in the hippocampus of SI C57BL/6J mice; the expression of the γ2 subunit was decreased whereas that of the α1 did not change. Patch-clamp recordings in dentate gyrus (DG) granule cells obtained from SI C57BL/6J mice revealed a greater enhancement of tonic currents induced by α-(4,5,6,7-tetrahydroisoxazolo[5,4-c] pyridin-3-ol (THIP) compared to that in control C57BL/6J mice. These neurochemical, molecular and functional changes observed in SI C57BL/6J mice were associated with an increased EtOH intake and EtOH preference. Nevertheless, the increase in EtOH consumption did not restore the reduction in hippocampal 3α,5α-TH PROG induced by SI. EtOH self-administration blocked the changes in gene expression of the α4 subunit but not those of the δ and γ2 subunits induced by SI. In addition, EtOH self-administration did not block the SI-induced changes in GABAAR-mediated tonic inhibition in hippocampal granule cells but increased the frequency of basal GABAergic sIPSCs in DG granule cells. We conclude that self-administration of EtOH selectively abolishes the increase of α4 subunit but not other neurochemical, molecular, and functional modifications induced by SI prolonged mild stress.

Maternal separation attenuates the effect of adolescent social isolation on HPA axis responsiveness in adult rats

Adverse early life experiences that occur during childhood and adolescence can have negative impacts on behavior later in life. The main goal of our work was to assess how the association between stressful experiences during neonatal and adolescent periods may influence stress responsiveness and brain plasticity in adult rats. Stressful experiences included maternal separation and social isolation at weaning. Three hours of separation from the pups (3-14 PND) significantly increased frequencies of maternal arched-back nursing and licking-grooming across the first two weeks postpartum. Separation also induced a long-lasting increase in dams blood levels of corticosterone. Maternal separation did not modify brain and plasma allopregnanolone and corticosterone levels in adult offspring, but they demonstrate partial recovery from the reduction induced by social isolation during adolescence. Moreover, the enhancement of corticosterone and allopregnanolone levels induced by foot shock stress in socially isolated animals that were subjected to maternal separation was markedly reduced with respect to that observed in animals that were just socially isolated. All experimental groups showed a significant reduction of BDNF and Arc protein expression in the hippocampus. However, the reduction of BDNF observed in animals that were maternally separated and subjected to social isolation was less significantly pronounced than in animals that were just socially isolated. The results sustained the mismatch hypothesis stating that aversive experiences early in life trigger adaptive processes, thereby rendering an individual to be better adapted to aversive challenges later in life.

Altered stress responsiveness and hypothalamic-pituitary-adrenal axis function in male rat offspring of socially isolated parents

Social isolation in male rats at weaning results in reduced basal levels of the neuroactive steroid 3α,5α‐tetrahydroprogesterone (3α,5α‐TH PROG) in the brain and plasma as well as increased anxiety‐like behavior. We now show that socially isolated female rats also manifest a reduced basal cerebrocortical concentration of 3α,5α‐TH PROG as well as an anxiety‐like profile in the elevated plus‐maze and Vogel conflict tests compared with group‐housed controls. In contrast, despite the fact that they were raised under normal conditions, adult male offspring of male and female rats subjected to social isolation before mating exhibited an increased basal cerebrocortical level of 3α,5α‐TH PROG but no difference in emotional reactivity compared with the offspring of group‐housed parents. These animals also showed an increased basal activity of the hypothalamic‐pituitary‐adrenal axis as well as reduced abundance of corticotropin‐releasing factor in the hypothalamus and of corticotropin‐releasing factor receptor type 1 in the pituitary. Moreover, negative feedback regulation of hypothalamic‐pituitary‐adrenal axis activity by glucocorticoid was enhanced in association with up‐regulation of glucocorticoid receptor expression in the hippocampus. There was also attenuation of corticosterone release induced by foot‐shock stress in the offspring of socially isolated parents. The increase in the brain concentration of 3α,5α‐TH PROG induced by acute stress was also blunted in these animals. Our results thus show that a stressful experience before mating can influence neuroendocrine signaling in the next generation.

P.1.014 Spatial memory and expression levels of BDNF and steroid hormone receptors in the hippocampus of socially isolated offspring

Purpose: Rats deprived of social contact with other rats at a young age experience a form of prolonged stress that leads to long-lasting alteration in their behaviour profile. This chronic stress paradigm results in changes of emotional state, hormonal secretion, abnormal reactivity to environmental and pharmacological stimuli and changes in GABAA receptor plasticity and function [1,2]. Moreover, isolated rats, although showed a down-regulation of both brain-derived neurotrophic factor (BDNF) and activityregulated cytoskeletal associated protein (Arc) in the hippocampus, demonstrated an improvement in memory performances in the Morris water maze (MWM) together with an increase in the expression of BDNF and Arc in the hippocampus on both days 1 and 5 and increased plasma level of corticosterone on the 5th day of MWM training [3]. To examine possible trans-generational effects on social isolated offspring we have investigated (1) basal plasma levels of corticosterone, (2) spatial memory performance in the MWM, (3) expression levels of BDNF and Arc in the hippocampus during the 1th and 5th days of MWM. Moreover, in the hippocampus of both isolated and isolated offspring we measured the expression levels of glucocorticoid (GR) and mineralcorticoid (MR) receptors. Methods:Male Sprague-Dawley rats were used. Steroid levels were quantified in plasma by RIA. The levels of BDNF, Arc, GR and MR proteins were determined by Western Blot analysis. The measurement of spatial memory was performed trough the MWM. Data were analyzed by one or two-way ANOVA followed by Newman-Keuls post-hoc test. Results: In social isolated offspring the plasma levels of corticosterone were significantly increased by 31% (p< 0.05) vs group-housed offspring (controls). In the MWM, social isolated offspring showed a decrease in latency to reach the platform during the days of training, and a decrease of the proximity measure to the target zone during the probe trial. Moreover, although the basal levels of hippocampal BDNF and Arc did not differ significantly between the two groups of animals, we found a significant increase of BDNF expression at the 5th day of MWM in isolated offspring vs the control animals (p< 0.05). Finally, the expression of hippocampal GR was significantly augmented in both isolated and isolated offspring (+41%, p< 0.05; +42%, p< 0.05, respectively), while the expression of MR was significantly increased only in the isolated rats (+35%, p< 0.05). Conclusions: Our results suggest that a stressful condition that have been experienced by the parents in early life time influence hormonal secretion, neuronal plasticity and cognitive performances in the offspring. Thus, isolated offspring showed an improvement in learning and spatial memory in MWM vs the control animals. Likewise, high circulating levels of corticosterone with high levels of GR expression in the hippocampus may play an important role in improving the cognitive performance of these animals. These data are in agreement with the increased expression of BDNF since it is involved in various forms of hippocampal learning.