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Background: Stress is a major risk factor for psychiatric disorders, particularly when occuring during the sensitive period of adolescence. We showed previously that male rats exposed to major stressors during prepuberty exhibit amygdala-driven loss of parvalbumin neurons in the ventral hippocampus, hippocampal hyperactivity, and an increase in ventral tegmental area (VTA) dopamine (DA) neuron activity. In contrast, we now find that female rats are resilient to the effects of prepubertal stress, but are uniquely sensitive to postpubertal (PostP) stress that impacts the system via a unique pathway involving the nucleus reuniens. Aims & Objectives: Using a combined stress paradigm, we examine the mechanism by which PostP stress in females activates medial VTA DA neuron activity, which are the DA neurons projecting to the affect-associated ventral nucleus accumbens. Method: Female rats were subjected to a combination of footshock/restraint stress during PostP (PD41-50). single-unit extracellular recordings of RE neurons 1-2 and 5-6 weeks were done after stress. RTN local field potentials were recorded in the RTN for at PD41, PD51, PD61, and PD85. Low (30-50Hz) and high gamma (60-120Hz) were analyzed. Immunohistochemistry analysis of PV/PNN content was evaluated in the posterior RTN one week after PostP stress (PD61). Another cohort of female rats received a retrograde inhibitory AAV-DREADDs virus (AAVrg-hSyn-hM4D(Gi)-mCherry, 500nL) at PD 31 followed by a cannula implantation in the nucleus reunions. Females were injected with Clozapine N-oxide (CNO, 1mM/200nL) in the nucleus reunions 15 min before stress during PD41-50. The rats were recorded for DA activity in the VTA during adulthood (PD>65). Results: Although female rats failed to show activation of the amygdala after PostP stress, another region that also projects to ventral hippocampal parvalbumin neurons, the nucleus reuniens, shows selective activation 1-2 weeks and 5-6 weeks after PostP stress only in females. This is accompanied by a loss of parvalbumin interneurons in the ventral hippocampus. This loss of hippocampal parvalbumin neurons leads to ventral hippocampal activation and DA neuron overdrive; an effect that is attenuated by inhibition of the reuniens-hippocampal pathway via inhibitory retrograde DREADD injection into the ventral hippocampus and CNO injection into the nucleus reuniens. One region that potently regulates primary thalamic nuclei, including the nucleus reuniens, is the reticular nucleus of the thalamus (RTN), a region consisting of parvalbumin-containing GABAergic neurons. We found that after postpubertal stress there is first an activation of low gamma oscillations one day after stress (consistent with parvalbumin neuron overdrive) followed one week later by a loss of parvalbumin neurons in the region of the RTN associated with the nucleus reuniens. Discussion & Conclusion: These data support that female rats that are resilient to prepubertal stress but show sensitivity to PostP stress that leads to parvalbumin loss in the RTN, decreased RTN inhibition of the nucleus reuniens, parvalbumin loss in the ventral hippocampus, and activation of the affect-related ventromedial striatum. These data are consistent with human studies, in which, females tend to be exposed more often to PostP stress in terms of e.g. sexual abuse, and are more susceptible to affect-related disorders. [ABSTRACT FROM AUTHOR]

Stress exposure can result in several proinflammatory alterations in the brain, including overexpression of the inducible isoform of nitric oxide synthase (iNOS) in the medial prefrontal cortex (mPFC). These changes may be involved in the development of many psychiatric conditions. However, it is unknown if iNOS in mPFC plays a significant role in stress-induced behavioral changes. The endocannabinoid (ECB) system is also influenced by stress. Its activation seems to be a counter regulatory mechanism to prevent or decrease the stress-mediated neuroinflammatory consequences. However, it is unclear if the ECB system and iNOS interact to influence stress consequences. This study aimed to test the hypothesis that the anti-stress effect of iNOS inhibition in mPFC involves the local ECB system, particularly the CB1 cannabinoid receptors. Male Wistar rats with guide cannula aimed at the mPFC were submitted to acute restraint stress (RS) for 2 h. In the following morning, rats received bilateral microinjections of vehicle, AM251 (CB1 antagonist; 100 pmol), and/or 1400W(iNOS selective inhibitor; 10-4, 10-3, or 10-2 nmol) into the prelimbic area of mPFC (PL-mPFC) before being tested in the elevated plus-maze (EPM). iNOS inhibition by 1400Wprevented the anxiogenic-like effect observed in animals submitted to RS. The drug did not promote behavior changes in naive animals, demonstrating a stress-dependent effect. The 1400W-anti-stress effect was prevented by local pretreatment with AM251. Our data suggest that iNOS inhibition may facilitate the local endocannabinoid signaling, attenuating stress effects. [ABSTRACT FROM AUTHOR]

Background: Inducible or neuronal nitric oxide synthase gene deletion increases or decreases anxiety-like behavior in mice, respectively. Since nitric oxide and endocannabinoids interact to modulate defensive behavior, the former effect could involve a compensatory increase in basal brain nitric oxide synthase activity and/or changes in the endocannabinoid system. Thus, we investigated the expression and extinction of contextual fear conditioning of inducible nitric oxide knockout mice and possible involvement of endocannabinoids in these responses. Methods: We evaluated the effects of a preferential neuronal nitric oxide synthase inhibitor, 7-nitroindazol, nitric oxide synthase activity, and mRNA changes of nitrergic and endocannabinoid systems components in the medial prefrontal cortex and hippocampus of wild-type and knockout mice. The effects of URB597, an inhibitor of the fatty acid amide hydrolase enzyme, which metabolizes the endocannabinoid anandamide, WIN55,212-2, a nonselective cannabinoid agonist, and AM281, a selective CB1 antagonist, on contextual fear conditioning were also evaluated. Results: Contextual fear conditioning expression was similar in wild-type and knockout mice, but the latter presented extinction deficits and increased basal nitric oxide synthase activity in the medial prefrontal cortex. 7-Nitroindazol decreased fear expression and facilitated extinction in wild-type and knockout mice. URB597 decreased fear expression in wild-type and facilitated extinction in knockout mice, whereas WIN55,212-2 and AM281 increased it in wild-type mice. Nonconditioned knockout mice showed changes in the mRNA expression of nitrergic and endocannabinoid system components in the medial prefrontal cortex and hippocampus that were modified by fear conditioning. Conclusion: These data reinforce the involvement of the nitric oxide and endocannabinoids (anandamide) in stress-related disorders and point to a deregulation of the endocannabinoid system in situations where nitric oxide signaling is increased. [ABSTRACT FROM AUTHOR]

Stress exposure can result in several proinflammatory alterations in the brain, including overexpression of the inducible isoform of nitric oxide synthase (iNOS) in the medial prefrontal cortex (mPFC). These changes may be involved in the development of many psychiatric conditions. However, it is unknown if iNOS in mPFC plays a significant role in stress-induced behavioral changes. The endocannabinoid (ECB) system is also influenced by stress. Its activation seems to be a counter regulatory mechanism to prevent or decrease the stress-mediated neuroinflammatory consequences. However, it is unclear if the ECB system and iNOS interact to influence stress consequences. This study aimed to test the hypothesis that the anti-stress effect of iNOS inhibition in mPFC involves the local ECB system, particularly the CB 1 cannabinoid receptors. Male Wistar rats with guide cannula aimed at the mPFC were submitted to acute restraint stress (RS) for 2 h. In the following morning, rats received bilateral microinjections of vehicle, AM251 (CB 1 antagonist; 100 pmol), and/or 1400W (iNOS selective inhibitor; 10 -4 , 10 -3 , or 10 -2 nmol) into the prelimbic area of mPFC (PL-mPFC) before being tested in the elevated plus-maze (EPM). iNOS inhibition by 1400W prevented the anxiogenic-like effect observed in animals submitted to RS. The drug did not promote behavior changes in naive animals, demonstrating a stress-dependent effect. The 1400W-anti-stress effect was prevented by local pretreatment with AM251. Our data suggest that iNOS inhibition may facilitate the local endocannabinoid signaling, attenuating stress effects., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Coelho, Vila-Verde, Sartim, Uliana, Braga, Guimarães and Lisboa.)