Adolescent chemogenetic activation of dopaminergic neurons leads to reversible decreases in amphetamine-induced stereotypic behavior.

Chronic perturbations of neuronal activity can evoke homeostatic and new setpoints for neurotransmission. Using chemogenetics to probe the relationship between neuronal cell types and behavior, we recently found reversible decreases in dopamine (DA) transmission, basal behavior, and amphetamine (AMPH) response following repeated stimulation of DA neurons in adult mice. It is unclear, however, whether altering DA neuronal activity via chemogenetics early in development leads to behavioral phenotypes that are reversible, as alterations of neuronal activity during developmentally sensitive periods might be expected to induce persistent effects on behavior. To examine the impact of developmental perturbation of DA neuron activity on basal and AMPH behavior, we expressed excitatory hM3D(Gq) in postnatal DA neurons in TH-Cre and WT mice. Basal and CNO- or AMPH-induced locomotion and stereotypy was evaluated in a longitudinal design, with clozapine N-oxide (CNO, 1.0 mg/kg) administered across adolescence (postnatal days 15-47). Repeated CNO administration did not impact basal behavior and only minimally reduced AMPH-induced hyperlocomotor response in adolescent TH-CrehM3Dq mice relative to WThM3Dq littermate controls. Following repeated CNO administration, however, AMPH-induced stereotypic behavior robustly decreased in adolescent TH-CrehM3Dq mice relative to controls. A two-month CNO washout period rescued the diminished AMPH-induced stereotypic behavior. Our findings indicate that the homeostatic compensations that take place in response to chronic hM3D(Gq) stimulation during adolescence are temporary and are dependent on ongoing chemogenetic stimulation.

Activation of pyramidal neurons in the infralimbic cortex alleviates LPS-induced depressive-like behavior in mice.

The infralimbic (IL) cortex dysfunction has been implicated in major depressive disorder (MDD), yet the precise cellular and molecular mechanisms remain poorly understood. In this study, we investigated the role of layer V pyramidal neurons in a mouse model of MDD induced by repeated lipopolysaccharide (LPS) administration. Our results demonstrate that three days of systemic LPS administration induced depressive-like behavior and upregulated mRNA levels of interleukin-1ß (IL-1ß), tumor necrosis factor-alpha (TNF-α), and transforming growth factor-ß (TGF-ß) in the IL cortex. Electrophysiological recordings revealed a significant decrease in the intrinsic excitability of layer V pyramidal neurons in the IL following systemic LPS exposure. Importantly, chemogenetic activation of IL pyramidal neurons ameliorated LPS-induced depressive-like behavior. Additionally, LPS administration significantly increased microglial activity in the IL, as evidenced by a greater number of Ionized calcium binding adaptor molecule-1 (IBA-1)-positive cells. Morphometric analysis further unveiled enlarged soma, decreased branch numbers, and shorter branch lengths of microglial cells in the IL cortex following LPS exposure. Moreover, the activation of pyramidal neurons by clozapine-N-oxide increased the microglia branch length but did not change branch number or cytosolic area. These results collectively suggest that targeted activation of pyramidal neurons in the IL cortex mitigates microglial response and ameliorates depressive-like behaviors induced by systemic LPS administration. Therefore, our findings offer potential therapeutic targets for the development of interventions aimed at alleviating depressive symptoms by modulating IL cortical circuitry and microglial activity.

Evaluation of the pharmacodynamic interaction effect of augmentation agents with clozapine in patients with treatment-resistant schizophrenia: A simulation study of clinical data.

INTRODUCTION: The aim of the study was to evaluate interaction effect of various augmentation strategies with clozapine in patients with Treatment-resistant schizophrenia. METHODS: Data was extracted for change in positive and negative syndrome scale (PANSS) or brief psychiatric rating scale (BPRS) scores for monotherapy with various antipsychotic agents alone and their combination with clozapine. Individual patient data was generated using simulation of data (factorial trial framework) from published clinical trials for sample sizes from eight to 400 to evaluate interaction effect through linear modeling. Dose equivalents were calculated, and best fit models were determined for simulated data. RESULTS: The polynomial model was found to be the best fit for the simulated data to determine interaction effect of combination. The clozapine augmentation with risperidone and ziprasidone was found to be antagonistic, whereas it was additive for haloperidol, aripiprazole, and quetiapine. A synergistic effect was observed for ECT combined with clozapine (Interaction effect: -7.62; p <0.001). A sample size of 250-300 may be sufficient to demonstrate a clinically significant interaction in future trials. CONCLUSION: Clozapine may be augmented with electroconvulsive therapy, leading to the enhancement of antipsychotic effect. Though some antipsychotics like aripiprazole demonstrate additive effects, they may also add to the adverse effects.

Prenatal stress alters transcription of NMDA-type glutamate receptors in the hippocampus.

Prenatal stress increases the risk of neurodevelopmental disorders. NMDA-type glutamate receptor (NMDAR) activity plays an important pathophysiological role in the cortico-hippocampal circuit in these disorders. We tested the hypothesis that transcription of NMDAR subunits is modified in the frontal cortex (FCx) and hippocampus after exposure to prenatal restraint stress (PRS) in mice. At 10 weeks of age, male PRS offspring (n = 20) and non-stressed controls (NS, n = 20) were treated with haloperidol (1 mg/kg), clozapine (5 mg/kg) or saline twice daily for 5 days, before measuring social approach (SOC). Saline-treated and haloperidol-treated PRS mice had reduced SOC relative to NS (P < 0.01), but clozapine-treated PRS mice had similar SOC to NS mice. These effects of PRS were associated with increased transcription of NMDAR subunits encoded by GRIN2A and GRIN2B genes in the hippocampus but not FCx. GRIN transcription in FCx correlated positively with SOC, but hippocampal GRIN transcription had negative correlation with SOC. The ratio of GRIN2A/GRIN2B transcription is known to increase during development but was lower in PRS mice. These results suggest that GRIN2A and GRIN2B transcript levels are modified in the hippocampus by PRS, leading to life-long deficits in social behavior. These data have some overlap with the molecular pathophysiology of schizophrenia. Similar to PRS in mice, schizophrenia, has been associated with social withdrawal, with increased GRIN2 expression in the hippocampus, and reduced GRIN2A/GRIN2B expression ratios in the hippocampus. These findings suggest that PRS in mice may have construct validity as a preclinical model for antipsychotic drug development.

Chemogenetic inactivation of the nucleus reuniens and its projections to the orbital cortex produce deficits on discrete measures of behavioral flexibility in the attentional set-shifting task.

The nucleus reuniens (RE) of the ventral midline thalamus is a critical node in the communication between the orbitomedial prefrontal cortex (OFC) and the hippocampus (HF). While RE has been shown to directly participate in memory-associated functions through its connections with the medial prefrontal cortex and HF, less is known regarding the role of RE in executive functioning. Here, we examined the involvement of RE and its projections to the orbital cortex (ORB) in attention and behavioral flexibility in male rats using the attentional set shifting task (AST). Rats expressing the hM4Di DREADD receptor in RE were implanted with indwelling cannulas in either RE or the ventromedial ORB to pharmacologically inhibit RE or its projections to the ORB with intracranial infusions of clozapine-N-oxide hydrochloride (CNO). Chemogenetic-induced suppression of RE resulted in impairments in reversal learning and set-shifting. This supports a vital role for RE in behavioral flexibility - or the ability to adapt behavior to changing reward or rule contingencies. Interestingly, CNO suppression of RE projections to the ventromedial ORB produced impairments in rule abstraction - or dissociable effects elicited with direct RE suppression. In summary, the present findings indicate that RE, mediated in part by actions on the ORB, serves a critical role in the flexible use of rules to drive goal directed behavior. The cognitive deficits of various neurological disorders with impaired communication between the HF and OFC, may be partly attributed to alterations of RE -- as an established intermediary between these cortical structures.

Ventral hippocampus mediates inter-trial responding in signaled active avoidance.

The hippocampus has a central role in regulating contextual processes in memory. We have shown that pharmacological inactivation of ventral hippocampus (VH) attenuates the context-dependence of signaled active avoidance (SAA) in rats. Here, we explore whether the VH mediates intertrial responses (ITRs), which are putative unreinforced avoidance responses that occur between trials. First, we examined whether VH inactivation would affect ITRs. Male rats underwent SAA training and subsequently received intra-VH infusions of saline or muscimol before retrieval tests in the training context. Rats that received muscimol performed significantly fewer ITRs, but equivalent avoidance responses, compared to controls. Next, we asked whether chemogenetic VH activation would increase ITR vigor. In male and female rats expressing excitatory (hM3Dq) DREADDs, systemic CNO administration produced a robust ITR increase that was not due to nonspecific locomotor effects. Then, we examined whether chemogenetic VH activation potentiated ITRs in an alternate (non-training) test context and found it did. Finally, to determine if context-US associations mediate ITRs, we exposed rats to the training context for three days after SAA training to extinguish the context. Rats submitted to context extinction did not show a reliable decrease in ITRs during a retrieval test, suggesting that context-US associations are not responsible for ITRs. Collectively, these results reveal an important role for the VH in context-dependent ITRs during SAA. Further work is required to explore the neural circuits and associative basis for these responses, which may be underlie pathological avoidance that occurs in humans after threat has passed.

Elevated intrinsic cortical curvature in treatment-resistant schizophrenia: Evidence of structural deformation in functional connectivity areas and comparison with alternate indices of structure.

BACKGROUND: Research suggests structural and connectivity abnormalities in patients with treatment-resistant schizophrenia (TRS) compared to first-line responders and healthy-controls. However, measures of these abnormalities are often influenced by external factors like nicotine and antipsychotics, limiting their clinical utility. Intrinsic-cortical-curvature (ICC) presents a millimetre-scale measure of brain gyrification, highly sensitive to schizophrenia differences, and associated with TRS-like traits in early stages of the disorder. Despite this evidence, ICC in TRS remains unexplored. This study investigates ICC as a marker for treatment resistance in TRS, alongside structural indices for comparison. METHODS: We assessed ICC in anterior cingulate, dorsolateral prefrontal, temporal, and parietal cortices of 38 first-line responders, 30 clozapine-resistant TRS, 37 clozapine-responsive TRS, and 52 healthy-controls. For comparative purposes, Fold and Curvature indices were also analyzed. RESULTS: Adjusting for age, sex, nicotine-use, and chlorpromazine equivalence, principal findings indicate ICC elevations in the left hemisphere dorsolateral prefrontal (p < 0.001, η2partial = 0.142) and temporal cortices (LH p = 0.007, η2partial = 0.060; RH p = 0.011, η2partial = 0.076) of both TRS groups, and left anterior cingulate cortex of clozapine-resistant TRS (p = 0.026, η2partial = 0.065), compared to healthy-controls. Elevations that correlated with reduced cognition (p = 0.001) and negative symptomology (p < 0.034) in clozapine-resistant TRS. Fold and Curvature indices only detected group differences in the right parietal cortex, showing interactions with age, sex, and nicotine use. ICC showed interactions with age. CONCLUSION: ICC elevations were found among patients with TRS, and correlated with symptom severity. ICCs relative independence from sex, nicotine-use, and antipsychotics, may support ICC's potential as a viable marker for TRS, though age interactions should be considered.

Chemogenetic inhibition of the lateral hypothalamus effectively reduces food intake in rats in a translational proof-of-concept study.

Despite the therapeutic potential of chemogenetics, the method lacks comprehensive preclinical validation, hindering its progression to human clinical trials. We aimed to validate a robust but simple in vivo efficacy assay in rats which could support chemogenetic drug discovery by providing a quick, simple and reliable animal model. Key methodological parameters such as adeno-associated virus (AAV) serotype, actuator drug, dose, and application routes were investigated by measuring the food-intake-reducing effect of chemogenetic inhibition of the lateral hypothalamus (LH) by hM4D(Gi) designer receptor stimulation. Subcutaneous deschloroclozapine in rats transfected with AAV9 resulted in a substantial reduction of food-intake, comparable to the efficacy of exenatide. We estimated that the effect of deschloroclozapine lasts 1-3 h post-administration. AAV5, oral administration of deschloroclozapine, and clozapine-N-oxide were also effective but with slightly less potency. The strongest effect on food-intake occurred within the first 30 min after re-feeding, suggesting this as the optimal experimental endpoint. This study demonstrates that general chemogenetic silencing of the LH can be utilized as an optimal, fast and reliable in vivo experimental model for conducting preclinical proof-of-concept studies in order to validate the in vivo effectiveness of novel chemogenetic treatments. We also hypothesize based on our results that universal LH silencing with existing and human translatable genetic neuroengineering techniques might be a viable strategy to affect food intake and influence obesity.

Neuroreceptor Inhibition by Clozapine Triggers Mitohormesis and Metabolic Reprogramming in Human Blood Cells.

The antipsychotic drug clozapine demonstrates superior efficacy in treatment-resistant schizophrenia, but its intracellular mode of action is not completely understood. Here, we analysed the effects of clozapine (2.5-20 µM) on metabolic fluxes, cell respiration, and intracellular ATP in human HL60 cells. Some results were confirmed in leukocytes of clozapine-treated patients. Neuroreceptor inhibition under clozapine reduced Akt activation with decreased glucose uptake, thereby inducing ER stress and the unfolded protein response (UPR). Metabolic profiling by liquid-chromatography/mass-spectrometry revealed downregulation of glycolysis and the pentose phosphate pathway, thereby saving glucose to keep the electron transport chain working. Mitochondrial respiration was dampened by upregulation of the F0F1-ATPase inhibitory factor 1 (IF1) leading to 30-40% lower oxygen consumption in HL60 cells. Blocking IF1 expression by cotreatment with epigallocatechin-3-gallate (EGCG) increased apoptosis of HL60 cells. Upregulation of the mitochondrial citrate carrier shifted excess citrate to the cytosol for use in lipogenesis and for storage as triacylglycerol in lipid droplets (LDs). Accordingly, clozapine-treated HL60 cells and leukocytes from clozapine-treated patients contain more LDs than untreated cells. Since mitochondrial disturbances are described in the pathophysiology of schizophrenia, clozapine-induced mitohormesis is an excellent way to escape energy deficits and improve cell survival.

Acute manipulation of Drd1 neurons in the prefrontal cortex bidirectionally regulates anxiety and depression-like behaviors.

BACKGROUND CONTEXT: The medial prefrontal cortex (mPFC) has been implicated in modulating anxiety and depression. Manipulation of Drd1 neurons in the mPFC resulted in variable neuronal activity and, consequently, strikingly different behaviors. The acute regulation of anxiety- and depression-like behaviors by Drd1 neurons, a major neuronal subtype in the mPFC, has not yet been investigated. PURPOSE: The purpose of this study was to investigate whether acute manipulation of Drd1 neurons in the mPFC affects anxiety- and depression-like behaviors. STUDY DESIGN: Male Drd1-Cre mice were injected with an adeno-associated virus (AAV) expressing hM3DGq or hM4DGi. Clozapine-n-oxide (CNO, 1 mg/kg, i.p.) was injected 30 min before the behavioral tests. METHODS: Male Drd1-Cre mice were injected with AAV-Ef1α-DIO-hM4DGi-mCherry-WPRE-pA, AAV-Ef1α-DIO-hM3DGq-mCherry-WPRE-pA or AAV-Ef1α-DIO-mCherry-WPRE-pA. Three weeks later, whole-cell recordings after CNO (5 µM) were applied to the bath were used to validate the functional expression of hM4DGi and hM3DGq. Four groups of mice underwent all the behavioral tests, and after each of the tests, the mice were allowed to rest for 3-4 days. CNO (1 mg/kg) was injected intraperitoneally 30 min before the behavior test. Anxiety-like behaviors were evaluated by the open field test (OFT), the elevated plus maze test (EPMT), and the novelty-suppressed feeding test (NSFT). Depression-like behaviors were evaluated by the sucrose preference test (SPT) and force swimming test (FST). For all experiments, coronal sections of the targeted brain area were used to confirm virus expression. RESULTS: Whole-cell recordings from brain slices demonstrated that infusions of CNO (5 µM) into mPFC slices dramatically increased the firing activity of hM3DGq-mCherry+ neurons and abolished the firing activity of hM4DGi-mCherry+ neurons. Acute chemogenetic activation of Drd1 neurons in the mPFC increased the time spent in the central area in the OFT, increased the time spent in the open arms in the EMPT, decreased the latency to bite the food in the NSFT, increased the sucrose preference in the SPT, and decreased the immobility time in the FST. Acute chemogenetic inhibition of Drd1 neurons in the mPFC decreased the time spent in the central area in the OFT, decreased the time spent in the open arms in the EMPT, increased the latency to bite the food in the NSFT, decreased the sucrose preference in the SPT, and increased the immobility time in the FST. CONCLUSIONS: The present study showed that acute activation of Drd1 neurons in the mPFC produced rapid anxiolytic- and antidepressant-like effects, and acute inhibition had the opposite effect, revealing that Drd1 neurons in the mPFC bidirectionally regulate anxiety- and depression-like behaviors. CLINICAL SIGNIFICANCE: The findings of the present study regarding the acute effects of stimulating Drd1 neurons in the mPFC on anxiety and depression suggest that Drd1 neurons in the mPFC are a focus for the treatment of anxiety disorders and depression.

The neural basis of resting-state fMRI functional connectivity in fronto-limbic circuits revealed by chemogenetic manipulation.

Measures of fMRI resting-state functional connectivity (rs-FC) are an essential tool for basic and clinical investigations of fronto-limbic circuits. Understanding the relationship between rs-FC and the underlying patterns of neural activity in these circuits is therefore vital. Here we introduced inhibitory designer receptors exclusively activated by designer drugs (DREADDs) into the amygdala of two male macaques. We evaluated the causal effect of activating the DREADD receptors on rs-FC and neural activity within circuits connecting amygdala and frontal cortex. Activating the inhibitory DREADD increased rs-FC between amygdala and ventrolateral prefrontal cortex. Neurophysiological recordings revealed that the DREADD-induced increase in fMRI rs-FC was associated with increased local field potential coherency in the alpha band (6.5-14.5 Hz) between amygdala and ventrolateral prefrontal cortex. Thus, our multi-modal approach reveals the specific signature of neuronal activity that underlies rs-FC in fronto-limbic circuits.

Clearing the Fog: A Review of Antipsychotics for Parkinson's-Related Hallucinations: A Focus on Pimavanserin, Quetiapine and Clozapine.

Parkinson's disease is a progressive neurodegenerative disorder characterized by motor and non-motor symptoms, including hallucinations. The use of antipsychotic medications is a common strategy to manage hallucinations associated with Parkinson's disease psychosis (PDP). However, careful consideration is necessary when selecting the most appropriate drug due to the potential risks associated with the available treatment options. Atypical antipsychotics (AAPs), such as Pimavanserin and Clozapine, have effectively controlled PDP symptoms. On the contrary, the support for utilizing quetiapine is not as substantial as other antipsychotics because research studies specifically investigating its application are still emerging and relatively recent. The broad mechanisms of action of AAPs, involving dopamine and serotonin receptors, provide improved outcomes and fewer side effects than typical antipsychotics. Conversely, other antipsychotics, including risperidone, olanzapine, aripiprazole, ziprasidone, and lurasidone, have been found to worsen motor symptoms and are generally not recommended for PDP. While AAPs offer favorable benefits, they are associated with specific adverse effects. Extrapyramidal symptoms, somnolence, hypotension, constipation, and cognitive impairment are commonly observed with AAP use. Clozapine, in particular, carries a risk of agranulocytosis, necessitating close monitoring of blood counts. Pimavanserin, a selective serotonin inverse agonist, avoids receptor-related side effects but has been linked to corrected QT (QTc) interval prolongation, while quetiapine has been reported to be associated with an increased risk of mortality. This review aims to analyze the benefits, risks, and mechanisms of action of antipsychotic medications to assist clinicians in making informed decisions and enhance patient care.

Antipsychotic drugs selectively decorrelate long-range interactions in deep cortical layers.

Psychosis is characterized by a diminished ability of the brain to distinguish externally driven activity patterns from self-generated activity patterns. Antipsychotic drugs are a class of small molecules with relatively broad binding affinity for a variety of neuromodulator receptors that, in humans, can prevent or ameliorate psychosis. How these drugs influence the function of cortical circuits, and in particular their ability to distinguish between externally and self-generated activity patterns, is still largely unclear. To have experimental control over self-generated sensory feedback, we used a virtual reality environment in which the coupling between movement and visual feedback can be altered. We then used widefield calcium imaging to determine the cell type-specific functional effects of antipsychotic drugs in mouse dorsal cortex under different conditions of visuomotor coupling. By comparing cell type-specific activation patterns between locomotion onsets that were experimentally coupled to self-generated visual feedback and locomotion onsets that were not coupled, we show that deep cortical layers were differentially activated in these two conditions. We then show that the antipsychotic drug clozapine disrupted visuomotor integration at locomotion onsets also primarily in deep cortical layers. Given that one of the key components of visuomotor integration in cortex is long-range cortico-cortical connections, we tested whether the effect of clozapine was detectable in the correlation structure of activity patterns across dorsal cortex. We found that clozapine as well as two other antipsychotic drugs, aripiprazole and haloperidol, resulted in a strong reduction in correlations of layer 5 activity between cortical areas and impaired the spread of visuomotor prediction errors generated in visual cortex. Our results are consistent with the interpretation that a major functional effect of antipsychotic drugs is a selective alteration of long-range layer 5-mediated communication.

Chemogenetic Manipulation of Amygdala Kappa Opioid Receptor Neurons Modulates Amygdala Neuronal Activity and Neuropathic Pain Behaviors.

Neuroplasticity in the central nucleus of the amygdala (CeA) plays a key role in the modulation of pain and its aversive component. The dynorphin/kappa opioid receptor (KOR) system in the amygdala is critical for averse-affective behaviors in pain conditions, but its mechanisms are not well understood. Here, we used chemogenetic manipulations of amygdala KOR-expressing neurons to analyze the behavioral consequences in a chronic neuropathic pain model. For the chemogenetic inhibition or activation of KOR neurons in the CeA, a Cre-inducible viral vector encoding Gi-DREADD (hM4Di) or Gq-DREADD (hM3Dq) was injected stereotaxically into the right CeA of transgenic KOR-Cre mice. The chemogenetic inhibition of KOR neurons expressing hM4Di with a selective DREADD actuator (deschloroclozapine, DCZ) in sham control mice significantly decreased inhibitory transmission, resulting in a shift of inhibition/excitation balance to promote excitation and induced pain behaviors. The chemogenetic activation of KOR neurons expressing hM3Dq with DCZ in neuropathic mice significantly increased inhibitory transmission, decreased excitability, and decreased neuropathic pain behaviors. These data suggest that amygdala KOR neurons modulate pain behaviors by exerting an inhibitory tone on downstream CeA neurons. Therefore, activation of these interneurons or blockade of inhibitory KOR signaling in these neurons could restore control of amygdala output and mitigate pain.

The Dopaminergic Cells in the Median Raphe Region Regulate Social Behavior in Male Mice.

According to previous studies, the median raphe region (MRR) is known to contribute significantly to social behavior. Besides serotonin, there have also been reports of a small population of dopaminergic neurons in this region. Dopamine is linked to reward and locomotion, but very little is known about its role in the MRR. To address that, we first confirmed the presence of dopaminergic cells in the MRR of mice (immunohistochemistry, RT-PCR), and then also in humans (RT-PCR) using healthy donor samples to prove translational relevance. Next, we used chemogenetic technology in mice containing the Cre enzyme under the promoter of the dopamine transporter. With the help of an adeno-associated virus, designer receptors exclusively activated by designer drugs (DREADDs) were expressed in the dopaminergic cells of the MRR to manipulate their activity. Four weeks later, we performed an extensive behavioral characterization 30 min after the injection of the artificial ligand (Clozapine-N-Oxide). Stimulation of the dopaminergic cells in the MRR decreased social interest without influencing aggression and with an increase in social discrimination. Additionally, inhibition of the same cells increased the friendly social behavior during social interaction test. No behavioral changes were detected in anxiety, memory or locomotion. All in all, dopaminergic cells were present in both the mouse and human samples from the MRR, and the manipulation of the dopaminergic neurons in the MRR elicited a specific social response.

Differential impact of intermittent versus continuous treatment with clozapine on fatty acid metabolism in the brain of an MK-801-induced mouse model of schizophrenia.

Continuous antipsychotic treatment is often recommended to prevent relapse in schizophrenia. However, the efficacy of antipsychotic treatment appears to diminish in patients with relapsed schizophrenia and the underlying mechanisms are still unknown. Moreover, though the findings are inconclusive, several recent studies suggest that intermittent versus continuous treatment may not significantly differ in recurrence risk and therapeutic efficacy but potentially reduce the drug dose and side effects. Notably, disturbances in fatty acid (FA) metabolism are linked to the onset/relapse of schizophrenia, and patients with multi-episode schizophrenia have been reported to have reduced FA biosynthesis. We thus utilized an MK-801-induced animal model of schizophrenia to evaluate whether two treatment strategies of clozapine would affect drug response and FA metabolism differently in the brain. Schizophrenia-related behaviors were assessed through open field test (OFT) and prepulse inhibition (PPI) test, and FA profiles of prefrontal cortex (PFC) and hippocampus were analyzed by gas chromatography-mass spectrometry. Additionally, we measured gene expression levels of enzymes involved in FA synthesis. Both intermittent and continuous clozapine treatment reversed hypermotion and deficits in PPI in mice. Continuous treatment decreased total polyunsaturated fatty acids (PUFAs), saturated fatty acids (SFAs) and FAs in the PFC, whereas the intermittent administration increased n-6 PUFAs, SFAs and FAs compared to continuous administration. Meanwhile, continuous treatment reduced the expression of Fads1 and Elovl2, while intermittent treatment significantly upregulated them. This study discloses the novel findings that there was no significant difference in clozapine efficacy between continuous and intermittent administration, but intermittent treatment showed certain protective effects on phospholipid metabolism in the PFC.

Continuous oral olanzapine or clozapine treatment initiated in adolescence has differential short- and long-term impacts on antipsychotic sensitivity than those initiated in adulthood.

One of the major discoveries in recent research on antipsychotic drugs is that antipsychotic treatment in adolescence could induce robust long-term alterations in antipsychotic sensitivity that persist into adulthood. These long-term impacts are likely influenced by various factors, including the "diseased" state of animals, sex, type of drugs, mode of drug administration, and age of treatment onset. In this study we compared the short- and long-term behavioral effects of 21-day continuous oral olanzapine (7.5 mg/kg/day) or clozapine (30.0 mg/kg/day) administration in heathy or maternal immune activated adolescent (33-53 days old) or adult (80-100 days old) rats of both sexes. We used a conditioned avoidance response model to assess the drug-induced alterations in antipsychotic sensitivity. Here, we report that while under the chronic drug treatment period, olanzapine progressively increased its suppression of avoidance responding over time, especially when treatment was initiated in adulthood. Clozapine's suppression depended on the age of drug exposure, with treatment initiated in adulthood showing a suppression while that initiated in adolescent did not. After a 17-day drug-free interval, in a drug challenge test, olanzapine treatment initiated in adolescence caused a decrease in drug sensitivity, as reflected by less avoidance suppression (a tolerance effect); whereas that initiated in adulthood appeared to cause an increase (more avoidance suppression, a sensitization effect). Clozapine treatments initiated in both adolescence and adulthood caused a similar tolerance effect. Our findings indicate that the same chronic antipsychotic treatment regimen initiated in adolescence or adulthood can have differential short- and long-term impacts on drug sensitivity.

Disrupting direct inputs from the dorsal subiculum to the granular retrosplenial cortex impairs flexible spatial memory in the rat.

In a changing environment, animals must process spatial signals in a flexible manner. The rat hippocampal formation projects directly upon the retrosplenial cortex, with most inputs arising from the dorsal subiculum and terminating in the granular retrosplenial cortex (area 29). The present study examined whether these same projections are required for spatial working memory and what happens when available spatial cues are altered. Consequently, injections of iDREADDs were made into the dorsal subiculum of rats. In a separate control group, GFP-expressing adeno-associated virus was injected into the dorsal subiculum. Both groups received intracerebral infusions within the retrosplenial cortex of clozapine, which in the iDREADDs rats should selectively disrupt the subiculum to retrosplenial projections. When tested on reinforced T-maze alternation, disruption of the subiculum to retrosplenial projections had no evident effect on the performance of those alternation trials when all spatial-cue types remained present and unchanged. However, the same iDREADDs manipulation impaired performance on all three alternation conditions when there was a conflict or selective removal of spatial cues. These findings reveal how the direct projections from the dorsal subiculum to the retrosplenial cortex support the flexible integration of different spatial cue types, helping the animal to adopt the spatial strategy that best meets current environmental demands.

Sex-Specific Effects of Long-Term Antipsychotic Drug Treatment on Adipocyte Tissue and the Crosstalk to Liver and Brain in Rats.

Antipsychotic drug (APD) medication can lead to metabolic dysfunctions and weight gain, which together increase morbidity and mortality. Metabolically active visceral adipose tissue (VAT) in particular plays a crucial role in the etiopathology of these metabolic dysregulations. Here, we studied the effect of 12 weeks of drug medication by daily oral feeding of clozapine and haloperidol on the perirenal fat tissue as part of VAT of male and female Sprague Dawley rats in the context of complex former investigations on brain, liver, and blood. Adipocyte area values were determined, as well as triglycerides, non-esterified fatty acids (NEFAs), glucose, glycogen, lactate, malondialdehyde equivalents, ferric iron and protein levels of Perilipin-A, hormone-sensitive-lipase (HSL), hepcidin, glucose transporter-4 (Glut-4) and insulin receptor-ß (IR-ß). We found increased adipocyte mass in males, with slightly higher adipocyte area values in both males and females under clozapine treatment. Triglycerides, NEFAs, glucose and oxidative stress in the medicated groups were unchanged or slightly decreased. In contrast to controls and haloperidol-medicated rats, perirenal adipocyte mass and serum leptin levels were not correlated under clozapine. Protein expressions of perilipin-A, Glut-4 and HSL were decreased under clozapine treatment. IR-ß expression changed sex-specifically in the clozapine-medicated groups associated with higher hepcidin levels in the perirenal adipose tissue of clozapine-treated females. Taken together, clozapine and haloperidol had a smaller effect than expected on perirenal adipose tissue. The perirenal adipose tissue shows only weak changes in lipid and glucose metabolism. The main changes can be seen in the proteins examined, and probably in their effect on liver metabolism.