Disruptive effects of repeated stress on basolateral amygdala neurons and fear behavior across the estrous cycle in rats.

Stress is a precipitating factor in depression and anxiety disorders. Patients with these disorders often show amygdala abnormalities. The basolateral amygdala (BLA) is integral in mood and emotion, and is sensitive to stress. While much is known about effects of stress on BLA neuron activity and morphology in males, less is known in females. We tested whether repeated stress exerts distinct effects on BLA in vivo neuronal activity and morphology of Golgi-stained BLA neurons [lateral (LAT) and basal (BA) nuclei] in adult female rats. Repeated restraint stress increased BLA neuronal firing and caused hypertrophy of BLA neurons in males, while it decreased LAT and BA neuronal firing and caused hypotrophy of neurons in the LAT of females. BLA neuronal activity and function, such as fear conditioning, shifts across the estrous cycle. Repeated stress disrupted this pattern of BLA activity and fear expression over the estrous cycle. The disruptive effects of stress on the pattern of BLA function across estrous may produce behavior that is non-optimal for a specific phase of the estrous cycle. The contrasting effects of stress may contribute to sex differences in the effects of stress on mood and psychiatric disorders.

Sex- and Age-dependent Effects of Orexin 1 Receptor Blockade on Open-Field Behavior and Neuronal Activity.

Adolescence is a sensitive and critical period in brain development where psychiatric disorders such as anxiety, depression and post-traumatic stress disorder are more likely to emerge following a stressful life event. Females are two times more likely to suffer from psychiatric disorders than males. Patients with these disorders show alterations in orexins (also called hypocretins), important neuropeptides that regulate arousal, wakefulness and the hypothalamic-pituitary-adrenal axis activity. Little is known on the role of orexins in mediating arousal behaviors in male and female rats during adolescence or adulthood. Here, we examine the influence of orexin 1 receptor blockade by SB334867 in open-field behavior in male and female rats during early adolescence (PND 31-33) or adulthood (PND 75-77). Animals were injected with 0 (vehicle), 1, 10, or 30 mg/kg SB334867 (i.p.). Thirty minutes later, they were placed in an open field, and behavior and neuronal activity (c-Fos) were assessed. In adolescent males, SB334867 significantly increased immobility in the 10 mg/kg group compared to vehicle. However, this increase in immobility in adolescent males was not observed in adolescent females. In contrast to adolescent males, adult males in the 10 mg/kg dose group showed the opposite effect on immobility compared to vehicle. These results indicate that 10 mg/kg dose of SB334867 has opposing effects in adolescent and adult males, but few effects in adolescent and adult females. Differences in functional networks between limbic regions may underlie these effects of orexin receptor blockade that are sex- and age-dependent in rats.

Electrical stimulation of the hippocampal fimbria facilitates neuronal nitric oxide synthase activity in the medial shell of the rat nucleus accumbens: Modulation by dopamine D1 and D2 receptor activation.

The medial shell region of the nucleus accumbens (msNAc) is a key center for the regulation of goal-directed behavior and is likely to be dysfunctional in neuropsychiatric disorders such as addiction, depression and schizophrenia. Nitric oxide (NO)-producing interneurons in the msNAc are potently modulated by dopamine (DA) and may play an important role in synaptic integration in msNAc networks. In this study, neuronal NO synthase (nNOS) activity was measured in anesthetized rats using amperometric microsensors implanted into the msNAc or via histochemical techniques. In amperometric studies, NO oxidation current was recorded prior to and during electrical stimulation of the ipsilateral fimbria. Fimbria stimulation elicited a frequency and intensity-dependent increase in msNAc NO efflux which was attenuated by systemic administration of the nNOS inhibitor NG-propyl-l-arginine. Parallel studies using NADPH-diaphorase histochemistry to assay nNOS activity produced highly complementary outcomes. Moreover, systemic administration of either a DA D1 receptor agonist or a DA D2 receptor antagonist potentiated nNOS activity in the msNAc elicited by fimbria stimulation. These observations demonstrate for the first time that NO synthesis in nNOS expressing interneurons in the msNAc is facilitated by robust activation of hippocampal afferents in a manner that is differentially modulated by DA D1 and D2 receptor activation.

Sex- and Estrus-Dependent Differences in Rat Basolateral Amygdala.

Depression and anxiety are diagnosed almost twice as often in women, and the symptomology differs in men and women and is sensitive to sex hormones. The basolateral amygdala (BLA) contributes to emotion-related behaviors that differ between males and females and across the reproductive cycle. This hints at sex- or estrus-dependent features of BLA function, about which very little is known. The purpose of this study was to test whether there are sex differences or estrous cyclicity in rat BLA physiology and to determine their mechanistic correlates. We found substantial sex differences in the activity of neurons in lateral nuclei (LAT) and basal nuclei (BA) of the BLA that were associated with greater excitatory synaptic input in females. We also found strong differences in the activity of LAT and BA neurons across the estrous cycle. These differences were associated with a shift in the inhibition-excitation balance such that LAT had relatively greater inhibition during proestrus which paralleled more rapid cued fear extinction. In contrast, BA had relatively greater inhibition during diestrus that paralleled more rapid contextual fear extinction. These results are the first to demonstrate sex differences in BLA neuronal activity and the impact of estrous cyclicity on these measures. The shift between LAT and BA predominance across the estrous cycle provides a simple construct for understanding the effects of the estrous cycle on BLA-dependent behaviors. These results provide a novel framework to understand the cyclicity of emotional memory and highlight the importance of considering ovarian cycle when studying the BLA of females.SIGNIFICANCE STATEMENT There are differences in emotional responses and many psychiatric symptoms between males and females. This may point to sex differences in limbic brain regions. Here we demonstrate sex differences in neuronal activity in one key limbic region, the basolateral amygdala (BLA), whose activity fluctuates across the estrous cycle due to a shift in the balance of inhibition and excitation across two BLA regions, the lateral and basal nuclei. By uncovering this push-pull shift between lateral and basal nuclei, these results help to explain disparate findings about the effects of biological sex and estrous cyclicity on emotion and provide a framework for understanding fluctuations in emotional memory and psychiatric symptoms.

Greater physiological and behavioral effects of interrupted stress pattern compared to daily restraint stress in rats.

Repeated stress can trigger a range of psychiatric disorders, including anxiety. The propensity to develop abnormal behaviors after repeated stress is related to the severity, frequency and number of stressors. However, the pattern of stress exposure may contribute to the impact of stress. In addition, the anxiogenic nature of repeated stress exposure can be moderated by the degree of coping that occurs, and can be reflected in homotypic habituation to the repeated stress. However, expectations are not clear when a pattern of stress presentation is utilized that diminishes habituation. The purpose of these experiments is to test whether interrupted stress exposure decreases homotypic habituation and leads to greater effects on anxiety-like behavior in adult male rats. We found that repeated interrupted restraint stress resulted in less overall homotypic habituation compared to repeated daily restraint stress. This was demonstrated by greater production of fecal boli and greater corticosterone response to restraint. Furthermore, interrupted restraint stress resulted in a lower body weight and greater adrenal gland weight than daily restraint stress, and greater anxiety-like behavior in the elevated plus maze. Control experiments demonstrated that these effects of the interrupted pattern could not be explained by differences in the total number of stress exposures, differences in the total number of days that the stress periods encompased, nor could it be explained as a result of only the stress exposures after an interruption from stress. These experiments demonstrate that the pattern of stress exposure is a significant determinant of the effects of repeated stress, and that interrupted stress exposure that decreases habituation can have larger effects than a greater number of daily stress exposures. Differences in the pattern of stress exposure are therefore an important factor to consider when predicting the severity of the effects of repeated stress on psychiatric disorders.

Inhibition of striatal soluble guanylyl cyclase-cGMP signaling reverses basal ganglia dysfunction and akinesia in experimental parkinsonism.

OBJECTIVE: There is clearly a necessity to identify novel non-dopaminergic mechanisms as new therapeutic targets for Parkinson's disease (PD). Among these, the soluble guanylyl cyclase (sGC)-cGMP signaling cascade is emerging as a promising candidate for second messenger-based therapies for the amelioration of PD symptoms. In the present study, we examined the utility of the selective sGC inhibitor 1H-[1], [2], [4] oxadiazolo-[4,3-a]quinoxalin-1-one (ODQ) for reversing basal ganglia dysfunction and akinesia in animal models of PD. METHODS: The utility of the selective sGC inhibitor ODQ for reversing biochemical, electrophysiological, histochemical, and behavioral correlates of experimental PD was performed in 6-OHDA-lesioned rats and mice chronically treated with MPTP. RESULTS: We found that one systemic administration of ODQ is sufficient to reverse the characteristic elevations in striatal cGMP levels, striatal output neuron activity, and metabolic activity in the subthalamic nucleus observed in 6-OHDA-lesioned rats. The latter outcome was reproduced after intrastriatal infusion of ODQ. Systemic administration of ODQ was also effective in improving deficits in forelimb akinesia induced by 6-OHDA and MPTP. INTERPRETATION: Pharmacological inhibition of the sGC-cGMP signaling pathway is a promising non-dopaminergic treatment strategy for restoring basal ganglia dysfunction and attenuating motor symptoms associated with PD.

Stepping test in mice: a reliable approach in determining forelimb akinesia in MPTP-induced Parkinsonism.

Currently existing behavioral measures for motor impairments in rodent models with bilateral dopamine depletion have demonstrated to be difficult to assess due to the degree of task complexity. There is clearly a need for a behavioral test that is simplistic in design and does not require the animal to learn a specific task, in particular for mice. Here we adapted the stepping test, originally designed for assessing asymmetric motor deficits in rats (Olsson, M., Nikkhah, G., Bentlage, C., Bjorklund, A., 1995. Forelimb akinesia in the rat Parkinson model: differential effects of dopamine agonists and nigral transplants as assessed by a new stepping test. J. Neurosci. 15, 3863-3875; Schallert, T., De Ryck, M., Whishaw, I.Q., Ramirez, V.D., Teitelbaum, P., 1979. Excessive bracing reactions and their control by atropine and l-DOPA in an animal analog of Parkinsonism. Exp. Neurol. 64, 33-43), into a mouse-friendly version for bilateral dopamine lesion induced by subacute MPTP injection. We found that MPTP-treated mice exhibit a significant and persistent reduction in the number of adjusting steps when compared to saline-treated animals. Typically, MPTP-induced stepping deficit becomes apparent by the fourth MPTP injection. The number of adjusting steps continues to decline throughout the injections, and by day 10 from the last MPTP injection, the stepping deficit observed is associated with approximately 65% TH positive cells loss in the SN. Importantly, L-DOPA administration significantly improved stepping performance in MPTP-treated mice. Thus, stepping test in mice is a reliable and simple behavioral measure for assessing forelimb akinesia induced by systemic MPTP.