Parkinson's-linked LRRK2-G2019S derails AMPAR trafficking, mobility, and composition in striatum with cell-type and subunit specificity.

Parkinson's disease (PD) is a multifactorial disease that affects multiple brain systems and circuits. While defined by motor symptoms caused by degeneration of brainstem dopamine neurons, debilitating non-motor abnormalities in fronto-striatal-based cognitive function are common, appear early, and are initially independent of dopamine. Young adult mice expressing the PD-associated G2019S missense mutation in Lrrk2 also exhibit deficits in fronto-striatal-based cognitive tasks. In mice and humans, cognitive functions require dynamic adjustments in glutamatergic synapse strength through cell-surface trafficking of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid-type glutamate receptors (AMPARs), but it is unknown how LRRK2 mutation impacts dynamic features of AMPAR trafficking in striatal projection neurons (SPNs). Here, we used Lrrk2G2019S knockin mice to show that surface AMPAR subunit stoichiometry is altered biochemically and functionally in mutant SPNs in dorsomedial striatum to favor the incorporation of GluA1 over GluA2. GluA1-containing AMPARs were resistant to internalization from the cell surface, leaving an excessive accumulation of GluA1 on the surface within and outside synapses. This negatively impacted trafficking dynamics that normally support synapse strengthening, as GluA1-containing AMPARs failed to increase at synapses in response to a potentiating stimulus and showed significantly reduced surface mobility. Surface GluA2-containing AMPARs were expressed at normal levels in synapses, indicating subunit-selective impairment. Abnormal surface accumulation of GluA1 was independent of PKA activity and was limited to D1R SPNs. Since LRRK2 mutation is thought to be part of a common PD pathogenic pathway, our data suggest that sustained, striatal cell-type specific changes in AMPAR composition and trafficking contribute to cognitive or other impairments associated with PD.

Estrous Cycle Mediates Midbrain Neuron Excitability Altering Social Behavior upon Stress.

The estrous cycle is a potent modulator of neuron physiology. In rodents, in vivo ventral tegmental area (VTA) dopamine (DA) activity has been shown to fluctuate across the estrous cycle. Although the behavioral effect of fluctuating sex steroids on the reward circuit is well studied in response to drugs of abuse, few studies have focused on the molecular adaptations in the context of stress and motivated social behaviors. We hypothesized that estradiol fluctuations across the estrous cycle acts on the dopaminergic activity of the VTA to alter excitability and stress response. We used whole-cell slice electrophysiology of VTA DA neurons in naturally cycling, adult female C57BL/6J mice to characterize the effects of the estrous cycle and the role of 17ß-estradiol on neuronal activity. We show that the estrous phase alters the effect of 17ß-estradiol on excitability in the VTA. Behaviorally, the estrous phase during a series of acute variable social stressors modulates subsequent reward-related behaviors. Pharmacological inhibition of estrogen receptors in the VTA before stress during diestrus mimics the stress susceptibility found during estrus, whereas increased potassium channel activity in the VTA before stress reverses stress susceptibility found during estrus as assessed by social interaction behavior. This study identifies one possible potassium channel mechanism underlying the increased DA activity during estrus and reveals estrogen-dependent changes in neuronal function. Our findings demonstrate that the estrous cycle and estrogen signaling changes the physiology of DA neurons resulting in behavioral differences when the reward circuit is challenged with stress.SIGNIFICANCE STATEMENT The activity of the ventral tegmental area encodes signals of stress and reward. Dopaminergic activity has been found to be regulated by both local synaptic inputs as well as inputs from other brain regions. Here, we provide evidence that cycling sex steroids also play a role in modulating stress sensitivity of dopaminergic reward behavior. Specifically, we reveal a correlation of ionic activity with estrous phase, which influences the behavioral response to stress. These findings shed new light on how estrous cycle may influence dopaminergic activity primarily during times of stress perturbation.

LRRK2 mutation alters behavioral, synaptic, and nonsynaptic adaptations to acute social stress.

Parkinson's disease (PD) risk is increased by stress and certain gene mutations, including the most prevalent PD-linked mutation LRRK2-G2019S. Both PD and stress increase risk for psychiatric symptoms, yet it is unclear how PD-risk genes alter neural circuitry in response to stress that may promote psychopathology. Here we show significant differences between adult G2019S knockin and wild-type (wt) mice in stress-induced behaviors, with an unexpected uncoupling of depression-like and hedonia-like responses in G2019S mice. Moreover, mutant spiny projection neurons in nucleus accumbens (NAc) lack an adaptive, stress-induced change in excitability displayed by wt neurons, and instead show stress-induced changes in synaptic properties that wt neurons lack. Some synaptic alterations in NAc are already evident early in postnatal life. Thus G2019S alters the magnitude and direction of behavioral responses to stress that may reflect unique modifications of adaptive plasticity in cells and circuits implicated in psychopathology in humans.NEW & NOTEWORTHY Depression is associated with Parkinson's disease (PD), and environmental stress is a risk factor for both. We investigated how LRRK2-G2019S PD mutation affects depression-like behaviors, synaptic function, and intrinsic neuronal excitability following stress. In response to stress, the mutation drives abnormal synaptic changes, prevents adaptive changes in intrinsic excitability, and leads to aberrant behaviors, thus defining new ways in which PD mutations derail adaptive plasticity in response to stress that may contribute to disease onset.

Sex-specific peripheral and central responses to stress-induced depression and treatment in a mouse model.

Major depressive disorder affects ~20% of the world population and is characterized by strong sexual dimorphism with females being two to three times more likely to develop this disorder. Previously, we demonstrated that a combination therapy with dihydrocaffeic acid and malvidin-glucoside to synergistically target peripheral inflammation and stress-induced synaptic maladaptation in the brain was effective in alleviating chronic social defeat stress (CSDS)-induced depression-like phenotype in male mice. Here, we test the combination therapy in a female CSDS model for depression and compared sex-specific responses to stress in the periphery and the central nervous system. Similar to male mice, the combination treatment is also effective in promoting resilience against the CSDS-induced depression-like behavior in female mice. However, there are sex-specific differences in peripheral immune responses and differential gene regulation in the prefrontal cortex to chronic stress and to the treatment. These data indicate that while therapeutic approaches to combat stress-related disorders may be effective in both sexes, the mechanisms underlying these effects differ, emphasizing the need for inclusion of both sexes in preclinical studies using animal models.

Parkinson's LRRK2-G2019S risk gene mutation drives sex-specific behavioral and cellular adaptations to chronic variable stress.

Anxiety is a psychiatric non-motor symptom of Parkinson's that can appear in the prodromal period, prior to significant loss of brainstem dopamine neurons and motor symptoms. Parkinson's-related anxiety affects females more than males, despite the greater prevalence of Parkinson's in males. How stress, anxiety and Parkinson's are related and the basis for a sex-specific impact of stress in Parkinson's are not clear. We addressed this using young adult male and female mice carrying a G2019S knockin mutation of leucine-rich repeat kinase 2 ( Lrrk2 G2019S ) and Lrrk2 WT control mice. In humans, LRRK2 G2019S significantly elevates the risk of late-onset Parkinson's. To assess within-sex differences between Lrrk2 G2019S and control mice in stress-induced anxiety-like behaviors in young adulthood, we used a within-subject design whereby Lrrk2 G2019S and Lrrk2 WT control mice underwent tests of anxiety-like behaviors before (baseline) and following a 28 day (d) variable stress paradigm. There were no differences in behavioral measures between genotypes in males or females at baseline, indicating that the mutation alone does not produce anxiety-like responses. Following chronic stress, male Lrrk2 G2019S mice were affected similarly to male wildtypes except for novelty-suppressed feeding, where stress had no impact on Lrrk2 G2019S mice while significantly increasing latency to feed in Lrrk2 WT control mice. Female Lrrk2 G2019S mice were impacted by chronic stress similarly to wildtype females across all behavioral measures. Subsequent post-stress analyses compared cFos immunolabeling-based cellular activity patterns across several stress-relevant brain regions. The density of cFos-activated neurons across brain regions in both male and female Lrrk2 G2019S mice was generally lower compared to stressed Lrrk2 WT mice, except for the nucleus accumbens of male Lrrk2 G2019S mice, where cFos-labeled cell density was significantly higher than all other groups. Together, these data suggest that the Lrrk2 G2019S mutation differentially impacts anxiety-like behavioral responses to chronic stress in males and females that may reflect sex-specific adaptations observed in circuit activation patterns in stress-related brain regions.

Non-Motor Symptoms of Parkinson's Disease: The Neurobiology of Early Psychiatric and Cognitive Dysfunction.

Parkinson's disease (PD) is a progressive neurodegenerative disorder that has been recognized for over 200 years by its clinically dominant motor system impairment. There are prominent non-motor symptoms as well, and among these, psychiatric symptoms of depression and anxiety and cognitive impairment are common and can appear earlier than motor symptoms. Although the neurobiology underlying these particular PD-associated non-motor symptoms is not completely understood, the identification of PARK genes that contribute to hereditary and sporadic PD has enabled genetic models in animals that, in turn, have fostered ever deepening analyses of cells, synapses, circuits, and behaviors relevant to non-motor psychiatric and cognitive symptoms of human PD. Moreover, while it has long been recognized that inflammation is a prominent component of PD, recent studies demonstrate that brain-immune signaling crosstalk has significant modulatory effects on brain cell and synaptic function in the context of psychiatric symptoms. This review provides a focused update on such progress in understanding the neurobiology of PD-related non-motor psychiatric and cognitive symptoms.

Parkinson's-linked LRRK2-G2019S derails AMPAR trafficking, mobility and composition in striatum with cell-type and subunit specificity.

Parkinson's (PD) is a multi-factorial disease that affects multiple brain systems and circuits. While defined by motor symptoms caused by degeneration of brainstem dopamine neurons, debilitating non-motor abnormalities in fronto-striatal based cognitive function are common, appear early and are initially independent of dopamine. Young adult mice expressing the PD-associated G2019S missense mutation in Lrrk2 also exhibit deficits in fronto-striatal-based cognitive tasks. In mice and humans, cognitive functions require dynamic adjustments in glutamatergic synapse strength through cell-surface trafficking of AMPA-type glutamate receptors (AMPARs), but it is unknown how LRRK2 mutation impacts dynamic features of AMPAR trafficking in striatal projection neurons (SPNs). Here, we used Lrrk2 G2019S knockin mice to show that surface AMPAR subunit stoichiometry is altered biochemically and functionally in mutant SPNs to favor incorporation of GluA1 over GluA2. GluA1-containing AMPARs were resistant to internalization from the cell surface, leaving an excessive accumulation of GluA1 on the surface within and outside synapses. This negatively impacted trafficking dynamics that normally support synapse strengthening, as GluA1-containing AMPARs failed to increase at synapses in response to a potentiating stimulus and showed significantly reduced surface mobility. Surface GluA2-containing AMPARs were expressed at normal levels in synapses, indicating subunit-selective impairment. Abnormal surface accumulation of GluA1 was independent of PKA activity and was limited to D 1 R SPNs. Since LRRK2 mutation is thought to be part of a common PD pathogenic pathway, our data suggest that sustained, striatal cell-type specific changes in AMPAR composition and trafficking contribute to cognitive or other impairments associated with PD. SIGNIFICANCE STATEMENT: Mutations in LRRK2 are common genetic risks for PD. Lrrk2 G2019S mice fail to exhibit long-term potentiation at corticostriatal synapses and show significant deficits in frontal-striatal based cognitive tasks. While LRRK2 has been implicated generally in protein trafficking, whether G2019S derails AMPAR trafficking at synapses on striatal neurons (SPNs) is unknown. We show that surface GluA1-AMPARs fail to internalize and instead accumulate excessively within and outside synapses. This effect is selective to D 1 R SPNs and negatively impacts synapse strengthening as GluA1-AMPARs fail to increase at the surface in response to potentiation and show limited surface mobility. Thus, LRRK2-G2019S narrows the effective range of plasticity mechanisms, supporting the idea that cognitive symptoms reflect an imbalance in AMPAR trafficking mechanisms within cell-type specific projections.

Development of prefrontal corticostriatal connectivity in mice.

Rational decision making is grounded in learning to associate actions with outcomes, a process that depends on projections from prefrontal cortex to dorsomedial striatum. Symptoms associated with a variety of human pathological conditions ranging from schizophrenia and autism to Huntington's and Parkinson's disease point toward functional deficits in this projection, but its development is not well understood, making it difficult to investigate how perturbations in development of this circuitry could contribute to pathophysiology. We applied a novel strategy based on Hotspot Analysis to assess the developmental progression of anatomical positioning of prefrontal cortex to striatal projections. Corticostriatal axonal territories established at P7 expand in concert with striatal growth but remain largely unchanged in positioning through adulthood, indicating they are generated by directed, targeted growth and not modified extensively by postnatal experience. Consistent with these findings, corticostriatal synaptogenesis increased steadily from P7 to P56, with no evidence for widescale pruning. As corticostriatal synapse density increased over late postnatal ages, the strength of evoked PFC input onto dorsomedial striatal projection neurons also increased, but spontaneous glutamatergic synaptic activity was stable. Based on its pattern of expression, we asked whether the adhesion protein, Cdh8, influenced this progression. In mice lacking Cdh8 in PFC corticostriatal projection neurons, axon terminal fields in dorsal striatum shifted ventrally. Corticostriatal synaptogenesis was unimpeded, but spontaneous EPSC frequency declined and mice failed to learn to associate an action with an outcome. Collectively these findings show that corticostriatal axons grow to their target zone and are restrained from an early age, do not undergo postnatal synapse pruning as the most dominant models predict, and that a relatively modest shift in terminal arbor positioning and synapse function has an outsized, negative impact on corticostriatal-dependent behavior.

Development and cadherin-mediated control of prefrontal corticostriatal projections in mice.

Action-outcome associations depend on prefrontal cortex (PFC) projections to the dorsal striatum. To assess how these projections form, we measured PFC axon patterning, synapse formation, and functional maturation in the postnatally developing mouse striatum. Using Hotspot analysis, we show that PFC axons form an adult-like pattern of clustered terminations in the first postnatal week that remains largely stable thereafter. PFC-striatal synaptic strength is adult-like by P21, while excitatory synapse density increases until adulthood. We then tested how the targeted deletion of a candidate adhesion/guidance protein, Cadherin-8 (Cdh8), from corticostriatal neurons regulates pathway development. Mutant mice showed diminished PFC axon targeting and reduced spontaneous glutamatergic synaptic activity in the dorsal striatum. They also exhibited impaired behavioral performance in action-outcome learning. The data show that PFC-striatal axons form striatal territories through an early, directed growth model and they highlight essential contributions of Cdh8 to the anatomical and functional features critical for the formation of action-outcome associations.

Chemokine receptor 5 signaling in PFC mediates stress susceptibility in female mice.

Chronic stress induces changes in the periphery and the central nervous system (CNS) that contribute to neuropathology and behavioral abnormalities associated with psychiatric disorders. In this study, we examined the impact of peripheral and central inflammation during chronic social defeat stress (CSDS) in female mice. Compared to male mice, we found that female mice exhibited heightened peripheral inflammatory response and identified C-C motif chemokine ligand 5 (CCL5), as a stress-susceptibility marker in females. Blocking CCL5 signaling in the periphery promoted resilience to CSDS. In the brain, stress-susceptible mice displayed increased expression of C-C chemokine receptor 5 (CCR5), a receptor for CCL5, in microglia in the prefrontal cortex (PFC). This upregulation was associated with microglia morphological changes, their increased migration to the blood vessels, and enhanced phagocytosis of synaptic components and vascular material. These changes coincided with neurophysiological alterations and impaired blood-brain barrier (BBB) integrity. By blocking CCR5 signaling specifically in the PFC were able to prevent stress-induced physiological changes and rescue social avoidance behavior. Our findings are the first to demonstrate that stress-mediated dysregulation of the CCL5-CCR5 axis triggers excessive phagocytosis of synaptic materials and neurovascular components by microglia, resulting in disruptions in neurotransmission, reduced BBB integrity, and increased stress susceptibility. Our study provides new insights into the role of cortical microglia in female stress susceptibility and suggests that the CCL5-CCR5 axis may serve as a novel sex-specific therapeutic target for treating psychiatric disorders in females.

Chronic food restriction enhances dopamine-mediated intracranial self-stimulation.

Dopamine-mediated reinforcement and behavioral adaptation is essential to survival. Here, we test the effects of food restriction on dopamine-mediated learning and reinforcement using optical intracranial self-stimulation (oICSS), an optogenetic version of conventional electrical ICSS (also known as brain stimulation reward, BSR). Using mouse genetic lines to express channelrhodopsin selectively in midbrain dopamine neurons, we demonstrate that genetically expressed channelrhodopsin can mediate optically evoked dopamine release and support self-stimulation in a lever-pressing paradigm. Using this midbrain dopamine oICSS preparation, we compare acquisition and rate of pressing in ad libitum versus food restricted mice. Food restriction facilitated both more rapid acquisition of self-stimulation behavior and higher rates of responding; reversing food status after acquisition modulated response vigor in already established behavior. These data suggest that food restriction enhances both the acquisition and expression of dopamine-reinforced self-stimulation responding. These data demonstrate the utility of oICSS for examining changes in reinforcement learning concomitant to neuroadaptations induced in dopamine signaling by experimental manipulations such as food restriction.

Prolonged epigenomic and synaptic plasticity alterations following single exposure to a psychedelic in mice.

Clinical evidence suggests that rapid and sustained antidepressant action can be attained with a single exposure to psychedelics. However, the biological substrates and key mediators of psychedelics' enduring action remain unknown. Here, we show that a single administration of the psychedelic DOI produces fast-acting effects on frontal cortex dendritic spine structure and acceleration of fear extinction via the 5-HT2A receptor. Additionally, a single dose of DOI leads to changes in chromatin organization, particularly at enhancer regions of genes involved in synaptic assembly that stretch for days after the psychedelic exposure. These DOI-induced alterations in the neuronal epigenome overlap with genetic loci associated with schizophrenia, depression, and attention deficit hyperactivity disorder. Together, these data support that epigenomic-driven changes in synaptic plasticity sustain psychedelics' long-lasting antidepressant action but also warn about potential substrate overlap with genetic risks for certain psychiatric conditions.

Cyfip1 Regulates SynGAP1 at Hippocampal Synapses.

In humans, copy number variations in CYFIP1 appear to have sweeping physiological and structural consequences in the brain, either producing or altering the severity of intellectual disability, autism, and schizophrenia. Independently, SynGAP1 haploinsufficiency produces intellectual disability and, frequently, autism. Cyfip1 inhibits protein translation and promotes actin polymerization, and SynGAP1 is a synaptically localized Ras/Rap GAP. While these proteins are clearly distinct, studies investigating their functions in mice have shown that each regulates the maturation of synapses in the hippocampus and haploinsufficiency for either produces an exaggerated form of mGluR-dependent long-term depression, suggesting that some signaling pathways converge. In this study, we examined how Cyfip1 haploinsufficiency impacts SynGAP1 levels and localization, as well as potential sites for mechanistic interaction in mouse hippocampus. The data show that synaptic, but not total, levels of SynGAP1 in Cyfip1 +/- mice were abnormally low during early postnatal development and in adults. This may be in response to a shift in the balance of kinases that activate SynGAP1 as levels of Cdk5 were reduced and those of activated CaMKII were maintained in Cyfip1 +/- mice compared to wild-type mice. Alternatively, this could reflect altered actin dynamics as Rac1 activity in Cyfip1 +/- hippocampus was boosted significantly compared to wild-type mice, and levels of synaptic F-actin were generally enhanced due in part to an increase in the activity of the WAVE regulatory complex. Decreased synaptic SynGAP1 coupled with a CaMKII-mediated bias toward Rap1 inactivation at synapses is also consistent with increased levels of synaptic GluA2, increased AMPA receptor-mediated responses to stimulation, and increased levels of synaptic mGluR1/5 compared to wild-type mice. Collectively, our data suggest that Cyfip1 regulates SynGAP1 and the two proteins work coordinately at synapses to appropriately direct actin polymerization and GAP activity.