Molecular and Epigenetic Mechanisms for the Complex Effects of Stress on Synaptic Physiology and Cognitive Functions.

Evidence over the past decades has found that stress, particularly through the corticosterone stress hormones, produces complex changes in glutamatergic signaling in prefrontal cortex, which leads to the alteration of cognitive processes medicated by this brain region. Interestingly, the effects of stress on glutamatergic transmission appear to be "U-shaped," depending upon the duration and severity of the stressor. These biphasic effects of acute vs chronic stress represent the adaptive vs maladaptive responses to stressful stimuli. Animal studies suggest that the stress-induced modulation of excitatory synaptic transmission involves changes in presynaptic glutamate release, postsynaptic glutamate receptor membrane trafficking and degradation, spine structure and cytoskeleton network, and epigenetic control of gene expression. This review will discuss current findings on the key molecules involved in the stress-induced regulation of prefrontal cortex synaptic physiology and prefrontal cortex-mediated functions. Understanding the molecular and epigenetic mechanisms that underlie the complex effects of stress will help to develop novel strategies to cope with stress-related mental disorders.

Synergistic regulation of glutamatergic transmission by serotonin and norepinephrine reuptake inhibitors in prefrontal cortical neurons.

The monoamine system in the prefrontal cortex has been implicated in various mental disorders and has been the major target of anxiolytics and antidepressants. Clinical studies show that serotonin and norepinephrine reuptake inhibitors (SNRIs) produce better therapeutic effects than single selective reuptake inhibitors, but the underlying mechanisms are largely unknown. Here, we found that low dose SNRIs, by acting on 5-HT(1A) and α2-adrenergic receptors, synergistically reduced AMPA receptor (AMPAR)-mediated excitatory postsynaptic currents and AMPAR surface expression in prefrontal cortex pyramidal neurons via a mechanism involving Rab5/dynamin-mediated endocytosis of AMPARs. The synergistic effect of SNRIs on AMPARs was blocked by inhibition of activator of G protein signaling 3, a G protein modulator that prevents reassociation of G(i) protein α subunit and prolongs the ßγ-mediated signaling pathway. Moreover, the depression of AMPAR-mediated excitatory postsynaptic currents by SNRIs required p38 kinase activity, which was increased by 5-HT(1A) and α2-adrenergic receptor co-activation in an activator of G protein signaling 3-dependent manner. These results have revealed a potential mechanism for the synergy between the serotonin and norepinephrine systems in the regulation of glutamatergic transmission in cortical neurons.

Restoration of glutamatergic transmission by dopamine D4 receptors in stressed animals.

The prefrontal cortex (PFC), a key brain region for cognitive and emotional processes, is highly regulated by dopaminergic inputs. The dopamine D4 receptor, which is enriched in PFC, has been implicated in mental disorders, such as attention deficit-hyperactivity disorder and schizophrenia. Recently we have found homeostatic regulation of AMPA receptor-mediated synaptic transmission in PFC pyramidal neurons by the D4 receptor, providing a potential mechanism for D4 in stabilizing cortical excitability. Because stress is tightly linked to adaptive and maladaptive changes associated with mental health and disorders, we examined the synaptic actions of D4 in stressed rats. We found that neural excitability was elevated by acute stress and dampened by repeated stress. D4 activation produced a potent reduction of excitatory transmission in acutely stressed animals and a marked increase of excitatory transmission in repeatedly stressed animals. These effects of D4 targeted GluA2-lacking AMPA receptors and relied on the bi-directional regulation of calcium/calmodulin kinase II activity. The restoration of PFC glutamatergic transmission in stress conditions may enable D4 receptors to serve as a synaptic stabilizer in normal and pathological conditions.

Assembly of a beta2-adrenergic receptor--GluR1 signalling complex for localized cAMP signalling.

Central noradrenergic signalling mediates arousal and facilitates learning through unknown molecular mechanisms. Here, we show that the beta(2)-adrenergic receptor (beta(2)AR), the trimeric G(s) protein, adenylyl cyclase, and PKA form a signalling complex with the AMPA-type glutamate receptor subunit GluR1, which is linked to the beta(2)AR through stargazin and PSD-95 and their homologues. Only GluR1 associated with the beta(2)AR is phosphorylated by PKA on beta(2)AR stimulation. Peptides that interfere with the beta(2)AR-GluR1 association prevent this phosphorylation of GluR1. This phosphorylation increases GluR1 surface expression at postsynaptic sites and amplitudes of EPSCs and mEPSCs in prefrontal cortex slices. Assembly of all proteins involved in the classic beta(2)AR-cAMP cascade into a supramolecular signalling complex and thus allows highly localized and selective regulation of one of its major target proteins.

Novel CHATogether family-centered mental health care in the post-pandemic era: a pilot case and evaluation.

BACKGROUND: The COVID-19 pandemic impacted children, adolescents, and their families, with significant psychosocial consequences. The prevalence of anxiety, depression, and self-injurious behaviors increased in our youth, as well as the number of suicide attempts and hospitalizations related to suicidal ideation. Additionally, parents' mental health saw increasing rates of depression, irritability, and alcohol use combined with worsening family function, child-parent connectedness, positive family expressiveness, and increases in family conflict. In light of these statistics, we created CHATogether (Compassionate Home, Action Together), a pilot family-centered intervention using multi-faceted psychotherapeutic approaches to improve familial communication and relational health between adolescents and their parents. This paper discusses the implementation of the CHATogether intervention at the Adolescent Intensive Outpatient Program (IOP), providing an example of the intervention through an in-depth pilot case, and evaluation of the program's acceptability and feasibility. METHODS: This paper describes a case in detail and evaluation from a total of 30 families that completed CHATogether in the initial pilot. Each family had 4-6 one-hour CHATogether sessions during their 6-week treatment course at the IOP. Before and after CHATogether, adolescents and their parents separately completed a questionnaire designed to explore their perceived family conflicts. After completion of the program, participants completed a brief quality improvement survey to assess their overall experience with CHATogether. In the reported case, the family completed Patient-Reported Outcomes Measurement Information System (PROMIS) depressive and anxiety symptoms scales, Conflict Behavior Questionnaires (CBQ), 9-item Concise Health Risk Tracking Self-Report (CHRT-SR9), and help-seeking attitude from adults during distress and suicide concerns. RESULTS: The pilot case showed a trend of improvement in reported depressive and anxiety symptoms, child-parent conflicts, subfactors of suicide risk including pessimism, helplessness, and despair, help-seeking acceptability from parents for suicide concerns, and the establishment of individualized family relationship goals. Preliminary feedback from participating families demonstrated positive effects on intra-family communication and improvement in the overall family dynamic. Adolescents (n = 30/30) and their parents (n = 30/30) rated "strongly agree" or "agree" that their families had benefited from CHATogether and welcomed participation in future program development. CONCLUSION: This study presents CHATogether as a novel family-centered intervention to address post-pandemic family mental health stress, especially when a family system was disrupted and negatively affected the mental health of children and adolescents. The intervention facilitated positive child-parent communication on a variety of topics, through tools such as emotional expression and help-seeking behavior. The reported pilot case and evaluation suggested CHATogether's acceptability and feasibility in a clinical context. We also provided quality improvement feedback to guide future studies in establishing the efficacy of CHATogether and other similar models of clinical family interventions.

Homeostatic regulation of glutamatergic transmission by dopamine D4 receptors.

Alterations of synaptic transmission have been considered a core feature of mental disorders; thus, we examined the role of dopamine D(4) receptors, which is highly implicated in attention-deficit hyperactivity disorder and schizophrenia, in regulating synaptic functions of prefrontal cortex, a brain region critical for cognitive and emotional processes. We found that D(4) stimulation caused a profound depression or potentiation of AMPA receptor-mediated excitatory synaptic transmission in prefrontal cortex pyramidal neurons when their activity was elevated or dampened, respectively, which was accompanied by a D(4)-induced decrease or increase of AMPARs at synapses. The dual effects of D(4) on AMPAR trafficking and function was dependent on the D(4)-mediated bidirectional regulation of CaMKII activity via coupling to distinct signaling pathways, which provides a unique mechanism for D(4) receptors to serve as a homeostatic synaptic factor to stabilize cortical excitability.

Navigating Intersectional Identities: Clinical Considerations for Working With LGBTQ Asian American Youth.

LGBTQ Asian American youth face unique challenges related to their marginalized identities. It is well documented that Asian Americans who need mental health treatment access care at lower rates than White populations.1 Although Asian cultural values are often cited as reasons for decreased help-seeking behavior, research suggests structural barriers including cost, lack of culturally tailored services, and lack of knowledge of available resources as greater contributors to these disparities.1 Asian Americans have also been subject to the "model minority" myth, the stereotype that the community is universally high achieving, rule following, and well adjusted. This false narrative contributes to negative mental health outcomes driven by racial discrimination and homogenizing the Asian American experience. This masks the diversity in mental health needs among Asian Americans. In addition, LGBTQ Asian Americans experience microaggressions, the perception of being "not queer enough," and racism from LGBTQ spaces that often primarily cater to a White population.2.

The novel antipsychotic drug lurasidone enhances N-methyl-D-aspartate receptor-mediated synaptic responses.

N-Methyl-D-aspartate (NMDA) receptor (NMDAR) hypofunction has been postulated to contribute to the cognitive deficit of schizophrenia. In this study, we examined the effect of lurasidone (Latuda; Dainippon Sumitomo Pharma Co. Ltd., Tokyo, Japan), a newly approved atypical antipsychotic drug (APD), on NMDAR synaptic function in rat frontal cortical pyramidal neurons. In vivo administration of lurasidone produced a significant and selective enhancement of NMDAR-mediated synaptic responses and surface expression of NR2A and NR2B subunits. Lurasidone has high affinity for serotonin 5-HT(1A), 5-HT(2A), and 5-HT(7) receptors and dopamine D(2) receptors. In vivo administration of the 5-HT(7) receptor antagonist (2R)-1-[(3-hydroxyphenyl)sulfonyl]-2 -(2-(4-methyl-1-piperidinyl)ethyl)pyrrolidine (SB-269970) mimicked the enhancing effect of lurasidone on NMDAR responses, whereas the D(2) receptor antagonist haloperidol failed to do so. Previous studies have found that short-term administration of lurasidone reverses the cognitive impairment induced by subchronic administration of phencyclidine (PCP), an NMDAR noncompetitive antagonist. In this study, we found that lurasidone, as well as the prototypical atypical APD clozapine, restored NMDAR-mediated synaptic responses to normal levels in the PCP model of schizophrenia. These results suggest that NMDAR is the potential key molecular target of lurasidone, possibility via antagonizing 5-HT(7) receptors, which is consistent with evidence that 5-HT(7) receptor antagonism contributes to cognitive enhancement by atypical APDs in patients with schizophrenia.

Cellular mechanisms for dopamine D4 receptor-induced homeostatic regulation of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors.

Aberrant dopamine D(4) receptor function has been implicated in mental illnesses, including schizophrenia and attention deficit-hyperactivity disorder. Recently we have found that D(4) receptor exerts an activity-dependent bi-directional regulation of AMPA receptor (AMPAR)-mediated synaptic currents in pyramidal neurons of prefrontal cortex (PFC) via the dual control of calcium/calmodulin kinase II (CaMKII) activity. In this study, we examined the signaling mechanisms downstream of CaMKII that govern the complex effects of D(4) on glutamatergic transmission. We found that in PFC neurons at high activity state, D(4) suppresses AMPAR responses by disrupting the kinesin motor-based transport of GluR2 along microtubules, which was accompanied by the D(4) reduction of microtubule stability via a mechanism dependent on CaMKII inhibition. On the other hand, in PFC neurons at the low activity state, D(4) potentiates AMPAR responses by facilitating synaptic targeting of GluR1 through the scaffold protein SAP97 via a mechanism dependent on CaMKII stimulation. Taken together, these results have identified distinct signaling mechanisms underlying the homeostatic regulation of glutamatergic transmission by D(4) receptors, which may be important for cognitive and emotional processes in which dopamine is involved.

Amyloid beta peptide-(1-42) induces internalization and degradation of beta2 adrenergic receptors in prefrontal cortical neurons.

Emerging evidence indicates that amyloid ß peptide (Aß) initially induces subtle alterations in synaptic function in Alzheimer disease. We have recently shown that Aß binds to ß(2) adrenergic receptor (ß(2)AR) and activates protein kinase A (PKA) signaling for glutamatergic regulation of synaptic activities. Here we show that in the cerebrums of mice expressing human familial mutant presenilin 1 and amyloid precursor protein genes, the levels of ß(2)AR are drastically reduced. Moreover, Aß induces internalization of transfected human ß(2)AR in fibroblasts and endogenous ß(2)AR in primary prefrontal cortical neurons. In fibroblasts, Aß treatment also induces transportation of ß(2)AR into lysosome, and prolonged Aß treatment causes ß(2)AR degradation. The Aß-induced ß(2)AR internalization requires the N terminus of the receptor containing the peptide binding sites and phosphorylation of ß(2)AR by G protein-coupled receptor kinase, not by PKA. However, the G protein-coupled receptor kinase phosphorylation of ß(2)AR and the receptor internalization are much slower than that induced by ßAR agonist isoproterenol. The Aß-induced ß(2)AR internalization is also dependent on adaptor protein arrestin 3 and GTPase dynamin, but not arrestin 2. Functionally, pretreatment of primary prefrontal cortical neurons with Aß induces desensitization of ß(2)AR, which leads to attenuated response to subsequent stimulation with isoproterenol, including decreased cAMP levels, PKA activities, PKA phosphorylation of serine 845 on α-amino-2,3-dihydro-5-methyl-3-oxo-4-isoxazolepropanoic acid (AMPA) receptor subunit 1 (GluR1), and AMPA receptor-mediated miniature excitatory postsynaptic currents. This study indicates that Aß induces ß(2)AR internalization and degradation leading to impairment of adrenergic and glutamatergic activities.

Impaired alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor trafficking and function by mutant huntingtin.

Emerging evidence from studies of Huntington disease (HD) pathophysiology suggests that huntingtin (htt) and its associated protein HAP1 participate in intracellular trafficking and synaptic function. However, it is largely unknown whether AMPA receptor trafficking, which is crucial for controlling the efficacy of synaptic excitation, is affected by the mutant huntingtin with polyglutamine expansion (polyQ-htt). In this study, we found that expressing polyQ-htt in neuronal cultures significantly decreased the amplitude and frequency of AMPAR-mediated miniature excitatory postsynaptic current (mEPSC), while expressing wild-type huntingtin (WT-htt) increased mEPSC. AMPAR-mediated synaptic transmission was also impaired in a transgenic mouse model of HD expressing polyQ-htt. The effect of polyQ-htt on mEPSC was mimicked by knockdown of HAP1 and occluded by the dominant negative HAP1. Moreover, we found that huntingtin affected mESPC via a mechanism depending on the kinesin motor protein, KIF5, which controls the transport of GluR2-containing AMPARs along microtubules in dendrites. The GluR2/KIF5/HAP1 complex was disrupted and dissociated from microtubules in the HD mouse model. Together, these data suggest that AMPAR trafficking and function is impaired by mutant huntingtin, presumably due to the interference of KIF5-mediated microtubule-based transport of AMPA receptors. The diminished strength of glutamatergic transmission could contribute to the deficits in movement control and cognitive processes in HD conditions.

Disrupted GABAAR trafficking and synaptic inhibition in a mouse model of Huntington's disease.

Growing evidence suggests that Huntington's disease (HD), a neurodegenerative movement disorder caused by the mutant huntingtin (htt) with an expanded polyglutamine (polyQ) repeat, is associated with the altered intracellular trafficking and synaptic function. GABA(A) receptors, the key determinant of the strength of synaptic inhibition, have been found to bind to the huntingtin associated protein 1 (HAP1). HAP1 serves as an adaptor linking GABA(A) receptors to the kinesin family motor protein 5 (KIF5), controlling the transport of GABA(A) receptors along microtubules in dendrites. In this study, we found that GABA(A)R-mediated synaptic transmission is significantly impaired in a transgenic mouse model of HD expressing polyQ-htt, which is accompanied by the diminished surface expression of GABA(A) receptors. Moreover, the GABA(A)R/HAP1/KIF5 complex is disrupted and dissociated from microtubules in the HD mouse model. These results suggest that GABA(A)R trafficking and function is impaired in HD, presumably due to the interference of KIF5-mediated microtubule-based transport of GABA(A) receptors. The diminished inhibitory synaptic efficacy could contribute to the loss of the excitatory/inhibitory balance, leading to increased neuronal excitotoxicity in HD.

Acute stress enhances glutamatergic transmission in prefrontal cortex and facilitates working memory.

The prefrontal cortex (PFC), a key brain region controlling cognition and emotion, is strongly influenced by stress. While chronic stress often produces detrimental effects on these measures, acute stress has been shown to enhance learning and memory, predominantly through the action of corticosteroid stress hormones. We used a combination of electrophysiological, biochemical, and behavioral approaches in an effort to identify the cellular targets of acute stress. We found that behavioral stressors in vivo cause a long-lasting potentiation of NMDAR- and AMPAR-mediated synaptic currents via glucocorticoid receptors (GRs) selectively in PFC pyramidal neurons. This effect is accompanied by increased surface expression of NMDAR and AMPAR subunits in acutely stressed animals. Furthermore, behavioral tests indicate that working memory, a key function relying on recurrent excitation within networks of PFC neurons, is enhanced by acute stress via a GR-dependent mechanism. These results have identified a form of long-term potentiation of synaptic transmission induced by natural stimuli in vivo, providing a potential molecular and cellular mechanism for the beneficial effects of acute stress on cognitive processes subserved by PFC.

Ubiquitin-dependent lysosomal targeting of GABA(A) receptors regulates neuronal inhibition.

The strength of synaptic inhibition depends partly on the number of GABA(A) receptors (GABA(A)Rs) found at synaptic sites. The trafficking of GABA(A)Rs within the endocytic pathway is a key determinant of surface GABA(A)R number and is altered in neuropathologies, such as cerebral ischemia. However, the molecular mechanisms and signaling pathways that regulate this trafficking are poorly understood. Here, we report the subunit specific lysosomal targeting of synaptic GABA(A)Rs. We demonstrate that the targeting of synaptic GABA(A)Rs into the degradation pathway is facilitated by ubiquitination of a motif within the intracellular domain of the gamma2 subunit. Blockade of lysosomal activity or disruption of the trafficking of ubiquitinated cargo to lysosomes specifically increases the efficacy of synaptic inhibition without altering excitatory currents. Moreover, mutation of the ubiquitination site within the gamma2 subunit retards the lysosomal targeting of GABA(A)Rs and is sufficient to block the loss of synaptic GABA(A)Rs after anoxic insult. Together, our results establish a previously unknown mechanism for influencing inhibitory transmission under normal and pathological conditions.

CHATogether: a novel digital program to promote Asian American Pacific Islander mental health in response to the COVID-19 pandemic.

BACKGROUND: In response to the COVID-19 pandemic and the associated rise in anti-Asian hate crimes, we developed the Compassionate Home, Action Together program, (CHATogether) to support the mental health of the Asian American and Pacific Islander (AAPI) community. CHATogether is a culturally informed and virtually delivered support program that harnesses the talents of AAPI teens, young adults, parents, and mental health professionals who share a commitment to serve their local communities. METHODS: Our objective was to identify the active components, optimal utilization, potential benefits, and pertinent limitations of the CHATogether program during the 3 years since its inception in 2019. By that time, the program had developed six distinct component arms: interactive theater, mental health education, research, peer support and community outreach, collaboration, and AAPI mentorship. To work towards this objective, we conducted a qualitative study using thematic analysis and an inductive approach based on grounded theory (GT), in which we analyzed anonymized transcripts of four focus groups, comprised of 20 program participants (11 females; 9 males). RESULTS: We developed a model of two overarching domains, each with three underlying themes: I. Individual stressors: (1) Family conflict; (2) Cultural identity; and (3) Pandemic impact; and II. Collective stressors: (1) Stigma related to mental health and illness; (2) Pandemic uncertainty; and (3) Xenophobia and societal polarization. Strengths of the CHATogether program include its role as a conduit toward AAPI connectedness and pride as well as purpose in building community. Through support and mentorship, the program cultivates a unique platform that promotes healing and resiliency in response to pandemic stressors and beyond. CONCLUSIONS: CHATogether creates a safe space for the AAPI community. Through its methods of storytelling and encouraging creativity, CHATogether facilitates the discussion of challenging topics specific to the AAPI community. Given the national mental health crisis that is further being exacerbated by the COVID-19 pandemic, a digital prevention program such as CHATogether holds promise towards providing access to mental health resources and supporting early help-seeking behaviors for individuals in the AAPI community.

The stress hormone corticosterone increases synaptic alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors via serum- and glucocorticoid-inducible kinase (SGK) regulation of the GDI-Rab4 complex.

Corticosterone, the major stress hormone, plays an important role in regulating neuronal functions of the limbic system, although the cellular targets and molecular mechanisms of corticosteroid signaling are largely unknown. Here we show that a short treatment of corticosterone significantly increases alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR)-mediated synaptic transmission and AMPAR membrane trafficking in pyramidal neurons of prefrontal cortex, a key region involved in cognition and emotion. This enhancing effect of corticosterone is through a mechanism dependent on Rab4, the small GTPase-controlling receptor recycling between early endosome and plasma membrane. Guanosine nucleotide dissociation inhibitor (GDI), which regulates the cycle of Rab proteins between membrane and cytosol, forms an increased complex with Rab4 after corticosterone treatment. Corticosterone also triggers an increased GDI phosphorylation at Ser-213 by the serum- and glucocorticoid-inducible kinase (SGK). Moreover, AMPAR synaptic currents and surface expression and their regulation by corticosterone are altered by mutating Ser-213 on GDI. These results suggest that corticosterone, via SGK phosphorylation of GDI at Ser-213, increases the formation of GDI-Rab4 complex, facilitating the functional cycle of Rab4 and Rab4-mediated recycling of AMPARs to the synaptic membrane. It provides a potential mechanism underlying the role of corticosteroid stress hormone in up-regulating excitatory synaptic efficacy in cortical neurons.

Dopamine D4 receptors regulate AMPA receptor trafficking and glutamatergic transmission in GABAergic interneurons of prefrontal cortex.

GABAergic interneurons in prefrontal cortex (PFC) play a critical role in cortical circuits by providing feedforward and feedback inhibition and synchronizing neuronal activity. Impairments in GABAergic inhibition to PFC pyramidal neurons have been implicated in the abnormal neural synchrony and working memory disturbances in schizophrenia. The dopamine D(4) receptor, which is strongly linked to neuropsychiatric disorders, such as attention deficit-hyperactivity disorder (ADHD) and schizophrenia, is highly expressed in PFC GABAergic interneurons, while the physiological role of D(4) in these interneurons is largely unknown. In this study, we found that D(4) activation caused a persistent suppression of AMPAR-mediated synaptic transmission in PFC interneurons. This effect of D(4) receptors on AMPAR-EPSC was via a mechanism dependent on actin/myosin V motor-based transport of AMPA receptors, which was regulated by cofilin, a major actin depolymerizing factor. Moreover, we demonstrated that the major cofilin-specific phosphatase Slingshot, which was activated by calcineurin downstream of D(4) signaling, was required for the D(4) regulation of glutamatergic transmission. Thus, D(4) receptors, by using the unique calcineurin/Slingshot/cofilin signaling mechanism, regulate actin dynamics and AMPAR trafficking in PFC GABAergic interneurons. It provides a potential mechanism for D(4) receptors to control the excitatory synaptic strength in local-circuit neurons and GABAergic inhibition in the PFC network, which may underlie the role of D(4) receptors in normal cognitive processes and mental disorders.

Regulation of AMPA receptor channels and synaptic plasticity by cofilin phosphatase Slingshot in cortical neurons.

Cofilin, the major actin depolymerizing factor, modulates actin dynamics that contribute to spine morphology, synaptic transmission and plasticity. Much evidence implicates the cofilin inactivation kinase LIMK in synaptic function, but little is known about the cofilin activation phosphatase Slingshot in this regard. In this study, we found that suppressing endogenous Slingshot with small RNA interference or function-blocking antibody led to a dramatic reduction of AMPA receptor-mediated excitatory postsynaptic currents (EPSCs) in cortical neurons. Perturbation of Slingshot function also diminished the ability to express synaptic plasticity. Inactivating cofilin or disturbing actin dynamics reduced AMPAR-EPSCs in a Slingshot-dependent manner. Moreover, surface GluR 1 and synaptic GluR2/3 clusters were reduced by Slingshot knockdown. Our data suggest that Slingshot plays a pivotal role in AMPAR trafficking and synaptic transmission by controlling actin cytoskeleton via cofilin activation.

Modulation of neuronal excitability by serotonin-NMDA interactions in prefrontal cortex.

Both serotonin and NMDA signaling in prefrontal cortex (PFC) are implicated in mental disorders, including depression and anxiety. To understand their potential contributions to PFC neuronal excitability, we examined the effect of co-activation of 5-HT and NMDA receptors on action potential firing elicited by depolarizing current injection in PFC pyramidal neurons. In the presence of NMDA, a low concentration of the 5-HT(1A) agonist 8-OH-DPAT substantially reduced the number of spikes, and a low concentration of the 5-HT(2A/C) agonist alpha-Me-5HT significantly enhanced it, while both agonists were ineffective when applied alone. The 8-OH-DPAT effect on firing was mediated by inhibition of protein kinase A (PKA), whereas the alpha-Me-5HT effect was mediated by activation of protein kinase C (PKC). Moreover, the extracellular signal-regulated kinase (ERK), a signaling molecule downstream of PKA and PKC, was involved in both 5-HT(1A) and 5-HT(2A/C) modulation of neuronal excitability. Biochemical evidence showed that 5-HT(1A) decreased, whereas 5-HT(2A/C) increased the activation of ERK in an NMDA-dependent manner. In animals exposed to acute stress, the enhancing effect of 5-HT(2A/C) on firing was lost, while the decreasing effect of 5-HT(1A) on firing was intact. Concomitantly, the effect of 5-HT(2A/C), but not 5-HT(1A), on ERK activation was abolished in stressed animals. Taken together, our results demonstrate that distinct 5-HT receptor subtypes, by interacting with NMDA receptors, differentially regulate PFC neuronal firing, and the complex effects of 5-HT receptors on excitability are selectively altered under stressful conditions, which are often associated with mental disorders.

Postsynaptic density-95 (PSD-95) and calcineurin control the sensitivity of N-methyl-D-aspartate receptors to calpain cleavage in cortical neurons.

The N-methyl-D-aspartate receptor (NMDAR) is a Ca(2+)-permeable glutamate receptor mediating many neuronal functions under normal and pathological conditions. Ca(2+) influx via NMDARs activates diverse intracellular targets, including Ca(2+)-dependent protease calpain. Biochemical studies suggest that NR2A and NR2B subunits of NMDARs are substrates of calpain. Our physiological data showed that calpain, activated by prolonged NMDA treatment (100 microM, 5 min) of cultured cortical neurons, irreversibly decreased the whole-cell currents mediated by extrasynaptic NMDARs. Animals exposed to transient forebrain ischemia, a condition that activates calpain, exhibited the reduced NMDAR current density and the lower full-length NR2A/B level in a calpain-dependent manner. Disruption of the association between NMDARs and the scaffolding protein postsynaptic density (PSD)-95 facilitated the calpain regulation of synaptic NMDAR responses and NR2 cleavage in cortical slices, whereas inhibition of calcineurin activity blocked the calpain effect on NMDAR currents and NR2 cleavage. Calpain-cleaved NR2B subunits were removed from the cell surface. Moreover, cell viability assays showed that calpain, by targeting NMDARs, provided a negative feedback to dampen neuronal excitability in excitotoxic conditions. These data suggest that calpain activation suppresses NMDAR function via proteolytic cleavage of NR2 subunits in vitro and in vivo, and the susceptibility of NMDARs to calpain cleavage is controlled by PSD-95 and calcineurin.