Brain adaptation to acute stress: effect of time, social buffering, and nicotinic cholinergic system.

Both social behavior and stress responses rely on the activity of the prefrontal cortex (PFC) and basolateral nucleus of the amygdala (BLA) and on cholinergic transmission. We previously showed in adult C57BL/6J (B6) mice that social interaction has a buffering effect on stress-related prefrontal activity, depending on the ß2-/- cholinergic nicotinic receptors (nAChRs, ß2-/- mice). The latency for this buffer to emerge being short, we question here whether the associated brain plasticity, as reflected by regional c-fos protein quantification and PFC-BLA functional connectivity, is modulated by time. Overall, we show that time normalized the stress-induced PFC hyperactivation in B6 mice and PFC hypo-activation in ß2-/- mice, with no effect on BLA. It also triggered a multitude of functional links between PFC subareas, and between PFC and BLA in B6 mice but not ß2-/- mice, showing a central role of nAChRs in this plasticity. Coupled with social interaction and time, stress led to novel and drastic diminution of functional connectivity within the PFC in both genotypes. Thus, time, emotional state, and social behavior induced dissociated effects on PFC and BLA activity and important cortico-cortical reorganizations. Both activity and plasticity were under the control of the ß2-nAChRs.

A multiscale analysis in CD38-/- mice unveils major prefrontal cortex dysfunctions.

Autism spectrum disorder (ASD) is characterized by early onset of behavioral and cognitive alterations. Low plasma levels of oxytocin (OT) have also been found in ASD patients; recently, a critical role for the enzyme CD38 in the regulation of OT release was demonstrated. CD38 is important in regulating several Ca2+-dependent pathways, but beyond its role in regulating OT secretion, it is not known whether a deficit in CD38 expression leads to functional modifications of the prefrontal cortex (PFC), a structure involved in social behavior. Here, we report that CD38-/- male mice show an abnormal cortex development, an excitation-inhibition balance shifted toward a higher excitation, and impaired synaptic plasticity in the PFC such as those observed in various mouse models of ASD. We also show that a lack of CD38 alters social behavior and emotional responses. Finally, examining neuromodulators known to control behavioral flexibility, we found elevated monoamine levels in the PFC of CD38-/- adult mice. Overall, our study unveiled major changes in PFC physiologic mechanisms and provides new evidence that the CD38-/- mouse could be a relevant model to study pathophysiological brain mechanisms of mental disorders such as ASD.-Martucci, L. L., Amar, M., Chaussenot, R., Benet, G., Bauer, O., de Zélicourt, A., Nosjean, A., Launay, J.-M., Callebert, J., Sebrié, C., Galione, A., Edeline, J.-M., de la Porte, S., Fossier, P., Granon, S., Vaillend, C., Cancela, J.-M., A multiscale analysis in CD38-/- mice unveils major prefrontal cortex dysfunctions.

Effects of water restriction on social behavior and 5-HT neurons density in the dorsal and median raphe nuclei in mice.

We explored here the hypothesis that temporary chronic water restriction in mice affects social behavior, via its action on the density of 5-HT neurons in dorsal and median raphe nuclei (DRN and MRN). For that, we submitted adult C57BL/6 J mice to mild and controlled temporary dehydration, i.e., 6 h of water access every 48 h for 15 days. We investigated their social behavior in a social interaction task known to allow free and reciprocal social contact. Results showed that temporary dehydration increases significantly time spent in social contact and social dominance. It also expands 5-HT neuron density within both DRN and MRN and the behavioral and neuronal plasticity were positively correlated. Our findings suggest that disturbance in 5-HT neurotransmission caused by temporary dehydration stress unbalances choice processes of animals in social context.

Dissociated features of social cognition altered in mouse models of schizophrenia: Focus on social dominance and acoustic communication.

Social and communication impairments are common features of psychiatric disorders. Animal models of schizophrenia display various social deficits due to difference in tests, mouse strains and drugs. Moreover, communication deficits have not been studied. Our objectives were to assess and compare three major features of social cognition in different mouse models of schizophrenia: interest for a social stimulus, organization and acceptance of social contact, and acoustic communication to question whether mouse models for schizophrenia with social dysfunction also exhibit vocal communication defects. To achieve these aims we treated acutely C57BL/6J mice either with MK-801 or ketamine and tested WT and microtubule-associated protein 6 -MAP6- KO mice in two complementary social tasks: the 3-chamber test which measures social motivation and the social interaction task -SIT- which relies on prefrontal cortex activity and measures the ability to organize and respond to a real interaction, and which promotes ultrasonic vocalizations. Our results reveal that schizophrenia models have intact interest for a social stimulus in the 3-chamber test. However, thanks to principal component analyses of social interaction data, we demonstrate that social motivation and the ability to act socially rely on distinct mechanisms in revealing a decrease in dominance and communication in pharmacological schizophrenia models along with social withdraw, classically observed in schizophrenia, in MK-801 model. In this latter model, some social parameters can be significantly improved by aripiprazole, an atypical antipsychotic. Our social protocol, combined with fine-tuned analysis, is expected to provide an innovative framework for testing future treatments in preclinical models. This article is part of the Special Issue entitled 'The neuropharmacology of social behavior: from bench to bedside'.

Stress-induced brain activation: buffering role of social behavior and neuronal nicotinic receptors.

Social behavior and stress responses both rely on activity in the prefrontal cortex (PFC) and amygdala. We previously reported that acute stress exposure impoverishes social repertoire and triggers behavioral rigidity, and that both effects are modulated by ß2-containing nicotinic receptors. We, therefore, hypothesized that the activity of brain regions associated with the integration of social cues will be modulated by stress exposure. We mapped the expression of c-fos protein in all subregions of the PFC and basolateral (BLA) and central (Ce) areas of the amygdala in C57BL/6J (B6) and ß2-/- mice. We show altered brain activity and differences in functional connectivity between the two genotypes after stress: the PFC and BLA were hyperactivated in B6 and hypo-activated in ß2-/- mice, showing that the impact of stress on brain activity and functional organization depends on the nicotinic system. We also show that the effects of the opportunity to explore a novel environment or interact socially after acute stress depended on genotype: exploration induced only marginal PFC activation in both genotypes relative to stress alone, excluding a major role for ß2 receptors in this process. However, social interaction following stress only activated the rostral and caudolateral areas of the PFC in B6 mice, while it induced a kindling of activation in all PFC and amygdalar areas in ß2-/- mice. These results indicate that acute stress triggers important PFC-amygdala plasticity, social interaction has a buffering role during stress-induced brain activation, and ß2 receptors contribute to the effects of social interaction under stress.

Social behaviors and acoustic vocalizations in different strains of mice.

Proposing a framework for the study of core functions is valuable for understanding how they are altered in multiple mental disorders involving prefrontal dysfunction, for understanding genetic influences and for testing therapeutic compounds. Social and communication disabilities are reported in several major psychiatric disorders, and social communication disorders also can occur independently. Being able to study social communication involving interactions and associated acoustic vocalizations in animal models is thus important. All rodents display extensive social behaviors, including interactions and acoustic vocalizations. It is therefore important to pinpoint potential genetic-related strain differences -and similarities- in social behavior and vocalization. One approach is to compare different mouse strains, and this may be useful in choosing which strains may be best suitable in modeling psychiatric disorders where social and communication deficits are core symptoms. We compared social behavior and ultrasonic acoustic vocalization profiles in males of four mouse strains (129S2/Sv, C57BL/6J, DBA/2, and CD-1) using a social interaction task that we previously showed to rely on prefrontal network activity. Our social interaction task promotes a high level of ultrasonic vocalization with both social and acoustic parameters, and further allows other measures of social behaviors. The duration of social contact, dominance and aggressiveness varied with the mouse strains. Only C57BL/6J mice showed no attacks, with social contact being highly affiliative, whereas others strains emitted aggressive attacks. C57BL/6J mice also exhibited a significantly higher rate of ultrasonic vocalizations (USV), especially during social interaction.

Trigemino-solitarii-facial pathway in rats.

This study was undertaken to identify premotor neurons in the nucleus tractus solitarii (NTS) serving as relay neurons between the sensory trigeminal complex (STC) and the facial motor nucleus in rats. Trigemino-solitarii connections were first investigated following injections of anterograde and/or retrograde (biotinylated dextran amine, biocytin, or gold-HRP) tracers in STC or NTS. Trigemino-solitarii neurons were abundant in the ventral and dorsal parts of the STC and of moderate density in its intermediate part. They project throughout the entire rostrocaudal extent of the NTS with a strong lateral preponderance. Solitarii-trigeminal neurons were observed mostly in the rostral and rostrolateral NTS. They mainly project to the ventral and dorsal parts of the spinal trigeminal nucleus but not to the principal nucleus. Additional neurons located in the middle NTS were found to project exclusively to the spinal trigeminal nucleus pars caudalis. No solitarii-trigeminal cells were observed in the caudal NTS. In addition, evidence was obtained of NTS retrogradely labeled neurons contacted by anterogradely labeled trigeminal terminals. Second, solitarii-facial projections were analyzed following injections of anterograde and retrograde tracers into the NTS and the facial nucleus, respectively. NTS neurons, except those of the rostrolateral part, reached the dorsal aspect of the facial nucleus. Finally, simultaneous injections of anterograde tracer in the STC and retrograde tracer in the facial nucleus gave retrogradely labeled neurons in the NTS receiving contacts from anterogradely labeled trigeminal boutons. Thus, the present data demonstrate for the first time the existence of a trigemino-solitarii-facial pathway. This could account for the involvement of the NTS in the control of orofacial motor behaviors.

5-HT2A receptors are expressed by catecholaminergic neurons in the rat nucleus tractus solitarii.

Within the nucleus tractus solitarii (NTS), serotonin (5-HT) exerts modulatory effects on central mechanisms controlling autonomic functions, notably through the stimulation of 5-HT2 receptors. Using double immunocytochemical labeling with specific anti-5-HT2A receptor antibodies and antibodies directed against the catecholamine (CA) synthesizing enzymes, i.e. tyrosine hydroxylase, dopamine beta-hydroxylase (DBH) and phenylethanolamine N-methyltransferase, we investigated whether 5-HT effects could be mediated by 5-HT2A receptors located on CA perikarya and/or processes within the NTS. A relatively high density of 5-HT2A immunoreactive processes was observed throughout the NTS. 5-HT2A neuronal perikarya were also found within the NTS except in its rostrolateral part. Double immunolabeling experiments revealed many 5-HT2A receptors on CA processes but only few 5-HT2A/TH and 5-HT2A/DBH perikarya. These data support the idea that, within the NTS, 5-HT2A-mediated control of autonomic functions involves CA neurons.

Involvement of NMDA receptors in the pressor response to microinjection of 5-HT3 agonist into the NTS of awake rats.

Previous studies have shown that the activation of 5-HT3 receptors in the nucleus tractus solitarii (NTS) increases the baseline mean arterial pressure (MAP). In the present study, we evaluated the possible involvement of NMDA receptors in this pressor response. Four days before the experiments, under tribromoethanol anesthesia, rats received two guide cannulas in the direction of the NTS, and 1 day before the experiments, under tribromoethanol anesthesia, the femoral artery was cannulated for pulsatile arterial pressure (PAP), MAP, and heart rate (HR) measurements. On the day of the experiments, 2-methyl-serotonin, a 5-HT3 agonist, was microinjected into the NTS after microinjection of saline or AP-5, a selective NMDA receptor antagonist. Microinjection of 2-methyl-serotonin (5 nmol/50 nl) into the NTS after the vehicle (saline) produced a significant increase in MAP (+ 20 +/- 5 mm Hg, n = 8) while microinjection of 2-methyl-serotonin after microinjection of AP-5 (10 nmol/50 nl) produced no change in baseline MAP (-1 +/- 3 mm Hg, n = 11). Microinjection of AP-5 into the NTS produced no significant changes in the baseline MAP and HR. The data show that the increase in MAP in response to microinjection of a 5-HT3 agonist into the NTS is dependent on NMDA receptors.

Acute stress in adulthood impoverishes social choices and triggers aggressiveness in preclinical models.

Adult C57BL/6J mice are known to exhibit high level of social flexibility while mice lacking the ß2 subunit of nicotinic receptors (ß2(-/-) mice) present social rigidity. We asked ourselves what would be the consequences of a restraint acute stress (45 min) on social interactions in adult mice of both genotypes, hence the contribution of neuronal nicotinic receptors in this process. We therefore dissected social interaction complexity of stressed and not stressed dyads of mice in a social interaction task. We also measured plasma corticosterone levels in our experimental conditions. We showed that a single stress exposure occurring in adulthood reduced and disorganized social interaction complexity in both C57BL/6J and ß2(-/-) mice. These stress-induced maladaptive social interactions involved alteration of distinct social categories and strategies in both genotypes, suggesting a dissociable impact of stress depending on the functioning of the cholinergic nicotinic system. In both genotypes, social behaviors under stress were coupled to aggressive reactions with no plasma corticosterone changes. Thus, aggressiveness appeared a general response independent of nicotinic function. We demonstrate here that a single stress exposure occurring in adulthood is sufficient to impoverish social interactions: stress impaired social flexibility in C57BL/6J mice whereas it reinforced ß2(-/-) mice behavioral rigidity.

Cholinergic differentiation of neural progenitors in adult mouse motor facial nucleus.

Environmental cues are critical determinants of the fate of neural progenitors (NPs) upon transplantation into the central nervous system. In the present study, we assessed the differentiation potential of NPs implanted in a cholinergic environment of the adult mouse brain. Neurospheres containing NPs issued from fetal ganglionic eminences of transgenic mice expressing the green fluorescent protein (GFP) were transplanted either inside or outside the mouse cholinergic facial motor nucleus. In some mice, a pre-degenerated nerve releasing trophic factors was grafted into this nucleus to favor NP survival and improve axonal growth into the graft. The fate of NPs was analyzed 6 to 9 days or 2 months post-transplantation by immunofluorescence under confocal microscopy. Transplanted NPs were observed both inside and outside the facial nucleus after 6 to 9 days, but almost exclusively inside after 2 months regardless of the presence of a pre-degenerated nerve. NPs expressed markers of undifferentiated cells, astrocytes, oligodendrocytes, neurons, or cholinergic cells. The cholinergic phenotype of NPs engrafted inside the facial nucleus increased with time and the presence of a pre-degenerated nerve. Large GFP cholinergic somata and abundant long cholinergic GFP axons projecting into the nerve graft were also observed. Our results show that NPs, isolated from fetal mouse brain and transplanted into the non-neurogenic environment of the adult mouse facial nucleus, differentiate into cholinergic cells capable to project axons. This environment and the nerve graft favored NP differentiation into cholinergic neurons.

Forced expression of the motor neuron determinant HB9 in neural stem cells affects neurogenesis.

In contrast to mouse embryonic stem cells and in spite of overlapping gene expression profiles, neural stem cells (NSCs) isolated from the embryonic spinal cord do not respond to physiological morphogenetic stimuli provided by Sonic hedgehog and retinoic acid and do not generate motor neurons upon differentiation. Transcription factors expressed in motor neuron progenitors during embryogenesis include Pax6, Ngn2, Nkx6.1 and Olig2, whose expression precedes that of factors specifying motor neuron fate, including HB9, Islet1 and LIM3. We showed that all these factors were present in neural progenitors derived from mouse ES cells, whereas NSCs derived from the rat embryonic spinal cord expressed neither HB9 nor Islet1 and contained low levels of Nkx6.1 and LIM3. We constructed a lentivirus vector to express HB9 and GFP in NSCs and examined the consequences of HB9 expression on other transcription factors and cell differentiation. Compared to cell expressing GFP alone, NSCs expressing GFP and HB9 cycled less rapidly, downregulated Pax6 and Ngn2 mRNA levels, produced higher proportions of neurons in vitro and lower numbers of neurons after transplantation in the spinal cord of recipient rats. Oligodendrocytic and astrocytic differentiations were not affected. HB9 expressing NSCs did not express Islet1 or upregulate LIM3. They neither responded to Sonic hedgehog and retinoic acid nor produced cholinergic neurons. We concluded that forced HB9 expression affected neurogenesis but was not sufficient to confer motor neuron fate to NSCs.

Serotonin transporter inhibitors protect against hypoxic pulmonary hypertension.

Pulmonary hypertension (PH) results from constriction and remodeling of pulmonary vessels. Serotonin contributes to both phenomena through different signaling pathways. The mitogenic effect of serotonin on pulmonary vascular smooth muscle cells is mediated by the serotonin transporter (5-hydroxytryptamine transporter [5-HTT]), whereas its constricting effect is mediated by 5-HT1B/1D and 5-HT2A receptors. Here, we investigated the respective roles of 5-HTT and 5-HT receptors on the development of chronic hypoxic PH in mice. During exposure to hypoxia (10% O2 for 2 weeks), the animals received one of the specific 5-HTT inhibitors citalopram and fluoxetine (10 mg/kg/day), the selective 5-HT1B/1D receptor antagonist GR127935 (2 and 10 mg/kg/day), or the 5-HT2A receptor antagonist ketanserin (2 mg/kg/day). Mice treated with the 5-HTT inhibitors showed less right ventricle hypertrophy (ratio of right ventricle/left ventricle + septum = 36.7 +/- 2.0% and 35.8 +/- 1.3% in citalopram- and fluoxetine-treated mice, respectively, vs. 41.5 +/- 1.5% in vehicle-treated mice) and less pulmonary vessel muscularization (p < 0.01) than those receiving the vehicle. Neither GR127935 nor ketanserin affected these parameters. These data indicate that 5-HTT plays a key role in hypoxia-induced pulmonary vascular remodeling. The effects of serotonin transporter inhibitors on PH in humans deserve investigation.