Prefrontal deep projection neurons enable cognitive flexibility via persistent feedback monitoring.

Cognitive flexibility, the ability to alter strategy according to changing stimulus-response-reward relationships, is critical for updating learned behavior. Attentional set-shifting, a test of cognitive flexibility, depends on the activity of prefrontal cortex (PFC). It remains unclear, however, what role PFC neurons play to support set-shifting. Using optogenetics and two-photon calcium imaging, we demonstrate that medial PFC activity does not bias sensorimotor responses during set-shifting, but rather enables set-shifting by encoding trial feedback information, a role it has been known to play in other contexts. Unexpectedly, the functional properties of PFC cells did not vary with their efferent projection targets. Instead, representations of trial feedback formed a topological gradient, with cells more strongly selective for feedback information located further from the pial surface, where afferent input from the anterior cingulate cortex was denser. These findings identify a critical role for deep PFC projection neurons in enabling set-shifting through behavioral feedback monitoring.

Early life adversity ablates sex differences in active versus passive threat responding in mice.

Early life adversity (ELA) heightens the risk for anxiety disorders (which are characterized by heightened fear and avoidance behaviors), with females being twice as likely as males to develop pathology. Pavlovian fear conditioning tasks have been used to study possible mechanisms supporting endophenotypes of pathology. Identification of sex and ELA selective effects on the nature of behavioral responding in these paradigms may provide a unique window into coping strategies in response to learned fear to guide more mechanistic studies. The goals of this study were two-fold; First, to test if male and female mice employed different coping strategies in response to threat learning using different conditioning parameters (low, medium, and high intensity foot shocks). Second, to test if ELA in the form of limited bedding and nesting (LBN) altered the behavioral response of mice to conditioning. Mice received 6 tone/foot-shock pairings at one of three different foot-shock intensities (0.35 mA; 0.57 mA; 0.7 mA). Freezing, darting, and foot-shock reactivity were measured across trials. During conditioning, control-reared female mice exhibited significantly higher rates of darting behavior compared to control males at nearly all shock intensities tested. LBN rearing decreased the proportion of darting females to levels observed in males. Thus, ELA in the form of LBN significantly diminished the recruitment of active versus passive coping strategies in female mice but did not generally change male responding. Additional work will be required to understand the neural basis of these behavioral effects. Findings extending from this work have the potential to shed light on how ELA impacts trajectories of regional brain development with implications for sex-selective risk for behavioral endophenotypes associated with pathology and possibly symptom presentation.

Revisiting the role of infralimbic cortex in fear extinction with optogenetics.

Previous rodent studies have implicated the infralimbic (IL) subregion of the medial prefrontal cortex in extinction of auditory fear conditioning. However, these studies used pharmacological inactivation or electrical stimulation techniques, which lack temporal precision and neuronal specificity. Here, we used an optogenetic approach to either activate (with channelrhodopsin) or silence (with halorhodopsin) glutamatergic IL neurons during conditioned tones delivered in one of two phases: extinction training or extinction retrieval. Activating IL neurons during extinction training reduced fear expression and strengthened extinction memory the following day. Silencing IL neurons during extinction training had no effect on within-session extinction, but impaired the retrieval of extinction the following day, indicating that IL activity during extinction tones is necessary for the formation of extinction memory. Surprisingly, however, silencing IL neurons optogenetically or pharmacologically during the retrieval of extinction 1 day or 1 week following extinction training had no effect. Our findings suggest that IL activity during extinction training likely facilitates storage of extinction in target structures, but contrary to current models, IL activity does not appear to be necessary for retrieval of extinction memory.

Early life adversity decreases pre-adolescent fear expression by accelerating amygdala PV cell development.

Early life adversity (ELA) is associated with increased risk for stress-related disorders later in life. The link between ELA and risk for psychopathology is well established but the developmental mechanisms remain unclear. Using a mouse model of resource insecurity, limited bedding (LB), we tested the effects of LB on the development of fear learning and neuronal structures involved in emotional regulation, the medial prefrontal cortex (mPFC) and basolateral amygdala (BLA). LB delayed the ability of peri-weanling (21 days old) mice to express, but not form, an auditory conditioned fear memory. LB accelerated the developmental emergence of parvalbumin (PV)-positive cells in the BLA and increased anatomical connections between PL and BLA. Fear expression in LB mice was rescued through optogenetic inactivation of PV-positive cells in the BLA. The current results provide a model of transiently blunted emotional reactivity in early development, with latent fear-associated memories emerging later in adolescence.

Early life stress leads to sex differences in development of depressive-like outcomes in a mouse model.

Childhood trauma and neglect influence emotional development and increase the risk for and severity of mental illness. Women have a heightened susceptibility to the effects of early life stress (ELS) and are twice as likely as men to develop debilitating, stress-associated disorders later in life, such as major depressive disorder (MDD). Until now, mouse models of depression have been largely unsuccessful at replicating the diverse symptomatology of this disease and the sex bias in vulnerability. From P4 to P11, a limited bedding model that leads to fragmented maternal care, was used to induce ELS. Early adolescent and young adult mice were tested on an array of assays to test for depressive-like behavior. This included our newly developed automated home cage behavioral recognition system, where the home cage behavior of ELS and control mice could be monitored over a continuous 5-10 day span. ELS females, but not males, exhibited depressive-like behaviors on traditional assays. These effects emerged during adolescence and became more severe in adulthood. Using the novel home cage video monitoring method, we identified robust and continuous markers of depressive-like pathology in ELS females that phenocopy many of the behavioral characteristics of depression in humans. ELS effects on home cage behavior were rapidly rescued by ketamine, a fast-acting antidepressant. Together, these findings highlight that limited bedding ELS (1) produces an early emerging, female-specific depressive phenotype that responds to a fast-acting antidepressant and (2) this model has the potential to inform sex-selective risk for the development of stress-induced mental illness.

Early Life Stress Delays Sexual Maturation in Female Mice.

In humans, some forms of early life stress (ELS) have been linked with precocious puberty, altered brain maturation, and increased risk for a variety of forms of pathology. Interestingly, not all forms of ELS have been found to equally impact these metrics of maturation. In recent work, we have found that ELS in the form of limited bedding (LB) from P4 to P11, was associated with precocious hippocampus maturation in males and increased risk for depressive-like pathology and attentional disturbance in female mice. Here, we sought to test whether ELS in the form of LB also impacted the timing of sexual maturation in female mice. To establish rate of somatic and sexual development, distinct cohorts of mice were tested for weight gain, timing of vaginal opening, and development of estrous cycling. ELS animals weighed significantly less than controls at every timepoint measured. Onset of vaginal opening was tracked from P21 to 40, and ELS was found to significantly delay the onset of vaginal opening. To test the impact of ELS on estrous cycle duration and regularity, vaginal cytology was assessed in independent groups of animals using either a continuous sampling (daily from P40 to P57) or random sampling approach (single swab at P35, P50, or P75). ELS did impact measures of estrous cycling, but these effects were dependent upon the sampling method used. We also tested the impact of ELS on anxiety-like behaviors over development and across the estrous cycle. We observed a developmental increase in anxiety-like behavior in control but not ELS mice. No effect of estrous cycle stage was found on anxiety-like behavior for either group of mice. Together these results provide evidence that ELS in the form of LB delays somatic and sexual development. Additional work will be required to determine the mechanism by which ELS impacts these measures, and if these effects are common to other models of ELS in rodents.

Acetylcholine Regulates Olfactory Perceptual Learning through Effects on Adult Neurogenesis.

Learning to perceptually discriminate between chemical signals in the environment (olfactory perceptual learning [OPL]) is critical for survival. Multiple mechanisms have been implicated in OPL, including modulation of neurogenesis and manipulation of cholinergic activity. However, whether these represent distinct processes regulating OPL or if cholinergic effects on OPL depend upon neurogenesis has remained an unresolved question. Using a combination of pharmacological and optogenetic approaches, cholinergic activity was shown to be both necessary and sufficient to drive OPL, and this process was dependent on the presence of newly born cells in the olfactory bulb (OB). This study is the first to directly demonstrate that cholinergic effects on OPL require adult OB neurogenesis.

Early life stress impairs contextual threat expression in female, but not male, mice.

Early life stress (ELS) is associated with altered processing of threat signals, and increased lifetime risk of anxiety and affective pathology, disorders that disproportionately affect females. We tested the impact of a limited bedding paradigm of ELS (from P4-11) on contextual threat learning, context memory, footshock sensitivity, and anxietylike behavior, in adult male and female mice. To examine contextual threat learning, mice conditioned by context/footshock association were tested 24 hr later for the context memory. To determine the effect of ELS on footshock sensitivity, a separate cohort of mice were exposed to footshocks of increasing intensity (0.06 to 0.40 mA) and behavioral responses (jump and audible vocalization) were assessed by observers blind to treatment condition, sex, and cycle stage. ELS impaired context memory in female, but not male, mice. ELS increased footshock-induced threshold to vocalize, but not to jump, in both sexes. In female mice, this effect was most apparent during estrus. Decreased body weight, indicative of higher stress incurred by an individual mouse, correlated with increased threshold to jump in both sexes reared in ELS, and to audibly vocalize in ELS females. As ELS effects on shock sensitivity were present in both sexes, the contextual recall deficit in females was not likely driven by changes in the salience of aversive footshocks. No effects on anxietylike behavior, as measured in the elevated plus maze (EPM), were observed. More work is needed to better understand the impact of ELS on both somatic and gonadal development, and their potential contribution to threat learning. (PsycINFO Database Record

Early Life Stress Drives Sex-Selective Impairment in Reversal Learning by Affecting Parvalbumin Interneurons in Orbitofrontal Cortex of Mice.

Poverty, displacement, and parental stress represent potent sources of early life stress (ELS). Stress disproportionately affects females, who are at increased risk for stress-related pathologies associated with cognitive impairment. Mechanisms underlying stress-associated cognitive impairment and enhanced risk of females remain unknown. Here, ELS is associated with impaired rule-reversal (RR) learning in females, but not males. Impaired performance was associated with decreased expression and density of interneurons expressing parvalbumin (PV+) in orbitofrontal cortex (OFC), but not other interneuron subtypes. Optogenetic silencing of PV+ interneuron activity in OFC of control mice phenocopied RR learning deficits observed in ELS females. Localization of reversal learning deficits to PV+ interneurons in OFC was confirmed by optogenetic studies in which neurons in medial prefrontal cortex (mPFC) were silenced and associated with select deficits in rule-shift learning. Sex-, cell-, and region-specific effects show altered PV+ interneuron development can be a driver of sex differences in cognitive dysfunction.

Early life stress leads to developmental and sex selective effects on performance in a novel object placement task.

Disruptions in early life care, including neglect, extreme poverty, and trauma, influence neural development and increase the risk for and severity of pathology. Significant sex disparities have been identified for affective pathology, with females having an increased risk of developing anxiety and depressive disorder. However, the effects of early life stress (ELS) on cognitive development have not been as well characterized, especially in reference to sex specific impacts of ELS on cognitive abilities over development. In mice, fragmented maternal care resulting from maternal bedding restriction, was used to induce ELS. The development of spatial abilities were tracked using a novel object placement (NOP) task at several different ages across early development (P21, P28, P38, P50, and P75). Male mice exposed to ELS showed significant impairments in the NOP task compared with control reared mice at all ages tested. In female mice, ELS led to impaired NOP performance immediately following weaning (P21) and during peri-adolescence (P38), but these effects did not persist into early adulthood. Prior work has implicated impaired hippocampus neurogenesis as a possible mediator of negative outcomes in ELS males. In the hippocampus of behaviorally naïve animals there was a significant decrease in expression of Ki-67 (proliferative marker) and doublecortin (DCX-immature cell marker) as mice aged, and a more rapid developmental decline in these markers in ELS reared mice. However, the effect of ELS dissipated by P28 and no main effect of sex were observed. Together these results indicate that ELS impacts the development of spatial abilities in both male and female mice and that these effects are more profound and lasting in males.