Buprenorphine prevents stress-induced blunting of nucleus accumbens dopamine response and approach behavior to food reward in mice.

Alterations to the mesolimbic dopamine (DA) system are thought to underlie dysfunctional reward processing in stress-related psychiatric disorders. Using in vivio microdialysis in awake freely moving mice, we assessed the effects of stress on the motivational and neurochemical correlates underlying conditioned approach behavior for palatable food in the non-deprived mouse. Mice trained to approach and consume food in a familiar environment exhibited a 30% increase in nucleus accumbens shell (AcbSh) extracellular dopamine levels coincident with approach towards and consumption of the food reward. This effect was not observed in mice that were presented with the food in an unfamiliar environment or were exposed for the first time and were region specific. The addition of an acute environmental stressor (bright light and novel scent) during food exposure decreased DA release and delayed approach to the food. The disruptive impact of acute novelty stress on DA levels and approach behavior was reversed in animals pretreated with buprenorphine, an opioid drug with antidepressant-like and anxiolytic effects. Together, these data indicate that exposure to mild stress reduces incentive drive to approach palatable food via alterations in AcbSh dopamine responsiveness to food reward. Moreover, they implicate the brain opioid system as a potential pharmacological target for counteracting behavioral and neurochemical elements associated with stress.

Overcoming resistance to single-agent therapy for oncogenic BRAF gene fusions via combinatorial targeting of MAPK and PI3K/mTOR signaling pathways.

Pediatric low-grade gliomas (PLGGs) are frequently associated with activating BRAF gene fusions, such as KIAA1549-BRAF, that aberrantly drive the mitogen activated protein kinase (MAPK) pathway. Although RAF inhibitors (RAFi) have been proven effective in BRAF-V600E mutant tumors, we have previously shown how the KIAA1549-BRAF fusion can be paradoxically activated by RAFi. While newer classes of RAFi, such as PLX8394, have now been shown to inhibit MAPK activation by KIAA1549-BRAF, we sought to identify alternative MAPK pathway targeting strategies using clinically relevant MEK inhibitors (MEKi), along with potential escape mechanisms of acquired resistance to single-agent MAPK pathway therapies. We demonstrate effectiveness of multiple MEKi against diverse BRAF-fusions with novel N-terminal partners, with trametinib being the most potent. However, resistance to MEKi or PLX8394 develops via increased RTK expression causing activation of PI3K/mTOR pathway in BRAF-fusion expressing resistant clones. To circumvent acquired resistance, we show potency of combinatorial targeting with trametinib and everolimus, an mTOR inhibitor (mTORi) against multiple BRAF-fusions. While single-agent mTORi and MEKi PLGG clinical trials are underway, our study provides preclinical rationales for using MEKi and mTORi combinatorial therapy to stave off or prevent emergent drug-resistance in BRAF-fusion driven PLGGs.

Corticotropin-releasing Factor in the Rat Dorsal Raphe Nucleus Promotes Different Forms of Behavioral Flexibility Depending on Social Stress History.

The stress-related neuropeptide, corticotropin-releasing factor (CRF) regulates the dorsal raphe nucleus-serotonin (DRN-5-HT) system during stress and this may underlie affective and cognitive dysfunctions that characterize stress-related psychiatric disorders. CRF acts on both CRF1 and CRF2 receptor subtypes in the DRN that exert opposing inhibitory and excitatory effects on DRN-5-HT neuronal activity and 5-HT forebrain release, respectively. The current study first assessed the cognitive effects of intra-DRN microinfusion of CRF or the selective CRF2 agonist, urocortin II in stress-naive rats on performance of an operant strategy set-shifting task that is mediated by the medial prefrontal cortex (mPFC). CRF (30 ng) facilitated strategy set-shifting performance, whereas higher doses of CRF and urocortin II that would interact with CRF2 were without effect, consistent with a CRF1-mediated action. This dose decreased 5-HT extracellular levels in the mPFC, further supporting a role for CRF1. The effects of CRF were then assessed in rats exposed to repeated social stress using the resident-intruder model. Repeated social stress shifted the CRF effect from facilitation of strategy set shifting to facilitation of reversal learning and this was most prominent in a subpopulation of rats that resist defeat. Notably, in this subpopulation of rats 5-HT neuronal responses to CRF have been demonstrated to shift from CRF1-mediated inhibition to CRF2-mediated excitation. Because 5-HT facilitates reversal learning, the present results suggest that stress-induced changes in the cellular effects of CRF in the DRN translate to changes in cognitive effects of CRF. Together, the results underscore the potential for stress history to shift cognitive processing through changes in CRF neurotransmission in the DRN and the association of this effect with coping strategy.

Effects of buprenorphine on behavioral tests for antidepressant and anxiolytic drugs in mice.

RATIONALE: Buprenorphine (BPN) has been shown to rapidly improve mood in treatment-resistant depressed patients in small clinical studies. However, BPN's effects in preclinical tests for mood and antidepressant efficacy are largely unexplored. OBJECTIVE: The current study examined the effects of BPN in the forced swim test (FST) and novelty-induced hypophagia (NIH) test as measures of antidepressant and anxiolytic-like effects in C57BL/6 J mice. Microdialysis was used to measure whether BPN engaged kappa-opioid receptor (KORs) in the nucleus accumbens shell (NAcSh) at a behaviorally active dose (0.25 mg/kg). METHODS: BPN was tested in the FST at both 30 min and 24 h post-administration. Also measured in the FST at 24 h post-administration were the KOR antagonist norbinaltorphimine (nor-BNI), the MOR agonist morphine and the reference antidepressant desipramine. The anxiolytic effects of BPN were examined in the NIH test 24 h after treatment. The effects of acute injection of BPN and the KOR agonist U50,488 were measured on extracellular dopamine (DA) levels in the NAcSh. RESULTS: BPN produced significant reductions in FST immobility without changing locomotor activity and reduced approach latencies in the novel environment of the NIH test when tested 24 h after treatment. Repeated daily BPN injections for 6 days did not produce tolerance to these behavioral effects. nor-BNI produced a similar antidepressant-like response in the FST 24 h post-injection but morphine and desipramine were ineffective. BPN (0.25 mg/kg) did not alter DA levels when given alone but prevented the KOR agonist U50,488 from reducing DA levels. CONCLUSIONS: Acute and subchronic treatment with BPN produced antidepressant and anxiolytic-like responses in mice at doses that engage KORs. These studies support the clinical evidence that BPN may be a novel rapid-acting antidepressant medication and provides rodent models for investigating associated neurochemical mechanisms.

D-serine deficiency attenuates the behavioral and cellular effects induced by the hallucinogenic 5-HT(2A) receptor agonist DOI.

Both the serotonin and glutamate systems have been implicated in the pathophysiology of schizophrenia, as well as in the mechanism of action of antipsychotic drugs. Psychedelic drugs act through the serotonin 2A receptor (5-HT2AR), and elicit a head-twitch response (HTR) in mice, which directly correlates to 5-HT2AR activation and is absent in 5-HT2AR knockout mice. The precise mechanism of this response remains unclear, but both an intrinsic cortico-cortical pathway and a thalamo-cortical pathway involving glutamate release have been proposed. Here, we used a genetic model of NMDAR hypofunction, the serine racemase knockout (SRKO) mouse, to explore the role of glutamatergic transmission in regulating 5-HT2AR-mediated cellular and behavioral responses. SRKO mice treated with the 5-HT2AR agonist (±)-2,5-dimethoxy-4-iodoamphetamine (DOI) showed a clearly diminished HTR and lower induction of c-fos mRNA. These altered functional responses in SRKO mice were not associated with changes in cortical or hippocampal 5-HT levels or in 5-HT2AR and metabotropic glutamate-2 receptor (mGluR2) mRNA and protein expression. Together, these findings suggest that D-serine-dependent NMDAR activity is involved in mediating the cellular and behavioral effects of 5-HT2AR activation.

Reversal of dopamine system dysfunction in response to high-fat diet.

OBJECTIVE: To test whether high-fat diet (HFD) decreases dopaminergic tone in reward regions of the brain and evaluate whether these changes reverse after removal of the HFD. DESIGN AND METHODS: Male and female mice were fed a 60% HFD for 12 weeks. An additional group was evaluated 4 weeks after removal of the HFD. These groups were compared with control fed, age-matched controls. Sucrose and saccharin preference was measured along with mRNA expression of dopamine (DA)-related genes by Real Time-quantitative PCR (RT-qPCR). DA and 3,4-dihydroxyphenylacetic acid (DOPAC) were measured using high-performance liquid chromatography. DNA methylation of the dopamine transporter (DAT) promoter was measured by methylated DNA immunoprecipitation and RT-qPCR. RESULTS: After chronic HFD, sucrose preference was reduced, and then normalized after removal of the HFD. Decreased expression of DA genes, decreased DA content and alterations in DAT promoter methylation, was observed. Importantly, response to HFD and the persistence of changes depended on sex and brain region. CONCLUSIONS: These data identify diminished DA tone after early-life chronic HFD with a complex pattern of reversal and persistence that varies by both sex and brain region. Central nervous system changes that did not reverse after HFD withdrawal may contribute to the difficulty in maintaining weight-loss after diet intervention.

Brain monoamines and antidepressant-like responses in MRL/MpJ versus C57BL/6J mice.

The MRL/MpJ mouse demonstrates enhanced wound healing and tissue regeneration and increased neurotrophic mobilization to chronic antidepressant drug treatments. This study compared brain monoamine systems between MRL/MpJ and C57BL/6J mice as a potential basis for strain differences after chronic antidepressant treatment. MRL/MpJ mice had significantly higher tissue levels of serotonin and dopamine in multiple brain regions. Microdialysis studies demonstrated that baseline levels of extracellular serotonin did not differ between strains. However, acute administration of the selective serotonin reuptake inhibitor citalopram produced an increase in extracellular serotonin in the ventral hippocampus of MRL/MpJ mice that was twice as large as achieved in C57BL/6J mice. The greater effects in MRL/MpJ mice on 5-HT levels were not maintained after local perfusion of citalopram, suggesting that mechanisms outside of the hippocampus were responsible for the greater effect of citalopram after systemic injection. The density of serotonin and norepinephrine transporters in the hippocampus was significantly higher in MRL/MpJ mice. In addition, the expression of 5-HT(1A) mRNA was lower in the hippocampus, 5-HT(1B) mRNA was higher in the hippocampus and brainstem and SERT mRNA was higher in the brain stem of MRL/MpJ mice. The exaggerated neurotransmitter release in MRL/MpJ mice was accompanied by reduced baseline immobility in the tail suspension test and a greater reduction of immobility produced by citalopram or the tricyclic antidepressant desipramine. These data suggest that differences in the response to acute and chronic antidepressant treatments between the two strains could be attributed to differences in serotonin or catecholamine transmission.

Akt1 deficiency in schizophrenia and impairment of hippocampal plasticity and function.

Genetic studies have associated deficient function of the serine/threonine kinase Akt1 with schizophrenia. This disorder is associated with developmental, structural, and functional abnormalities of the hippocampus that could be traced to abnormal Akt1 function. To establish a closer connection between Akt1 and hippocampal function, mice with a selective deletion of Akt1 (Akt1(-/-) mice) were examined for physiological and behavioral outcomes dependent on the hippocampus and associated with schizophrenia. Genetic deletion of Akt1 was associated with both impaired proliferative capacity of adult-born hippocampal progenitors and hippocampal long-term potentiation, indicating deficient functions of this brain region associated with neuroplasticity. Moreover, Akt1(-/-) mice demonstrated impairments in contextual fear conditioning and recall of spatial learning, behaviors known to selectively involve the hippocampus. Akt1(-/-) mice also showed reduced prepulse inhibition of the acoustic startle response, a sensorimotor gating response that is perturbed in schizophrenia. Postmortem tissue samples from patients with schizophrenia showed significant reductions of phosphorylated Akt levels in hilar neurons of the dentate gyrus, the neurogenic zone of the hippocampus. Taken together, these results implicate the Akt1 isoform in regulating hippocampal neuroplasticity and cognition and in contributing to the etiology of schizophrenia.

Fluoxetine treatment induces dose dependent alterations in depression associated behavior and neural plasticity in female mice.

Antidepressant-induced increases in neurogenesis and neurotrophin mobilization in rodents and primates are proposed to be necessary for behavioral efficacy. The current study examines the relationship between the effects of fluoxetine treatment on behavior, cell proliferation and the neurotrophin BDNF in females. Female MRL/MpJ mice were treated acutely (5 and 10mg/kg) or chronically (2.5, 5 and 10mg/kg b.i.d.) with fluoxetine and tested in the tail suspension test (TST) and or novelty-induced hypophagia test (NIH), respectively. Mice treated chronically with fluoxetine received 4 (100mg/kg) injections of 5-bromo-2'-deoxyuridine (BrdU) on the last 4 days of treatment to measure DNA synthesis. The other half of the hippocampus and the frontal cortex was removed and examined for BDNF levels. Fluoxetine treatment decreased immobility in the TST and latency to eat in the NIH test, but only the highest dose of fluoxetine significantly altered behavior in both tests. Chronic treatment with 5 and 10mg/kg of fluoxetine significantly increased cell proliferation and BDNF levels in the hippocampus. Only chronic treatment with the highest of fluoxetine increased BDNF levels in the frontal cortex. Behavioral measures in the NIH test correlated with BDNF levels in the frontal cortex but not in the hippocampus or with cell proliferation in the hippocampus. These data suggest that females require high doses of fluoxetine for behavioral efficacy regardless of elevations of neurogenesis and BDNF mobilization in the hippocampus. Elevations in BDNF levels in the frontal cortex are related to the behavioral efficacy of fluoxetine.

Sex-specific effects of chronic fluoxetine treatment on neuroplasticity and pharmacokinetics in mice.

Neurogenesis is a mechanism through which antidepressants may produce therapeutic effects. There is a dearth of information regarding the effects of antidepressants on neurogenesis and neurotrophin mobilization in females. This study examined sex differences in the alteration of cell proliferation and survival in multiple regions of the brain. Additional experiments examined brain-derived neurotrophic factor (BDNF) levels and pharmacokinetics of fluoxetine to determine whether they mediate sex differences. MRL/MpJ mice were treated with fluoxetine (5 and 10 mg/kg b.i.d.) for 21 days and received injections of 5-bromo-2'-deoxyuridine (200 mg/kg) to measure DNA synthesis. In the hippocampus, fluoxetine increased cell proliferation at both doses; females treated with 10 mg/kg produced more new cells than males. Fluoxetine did not alter survival in males, but 10 mg/kg reduced survival in females. In the frontal cortex, fluoxetine increased cell proliferation and survival in males treated with 10 mg/kg. In the cerebellum and amygdala, 10 mg/kg fluoxetine increased cell proliferation in both sexes but did not alter the incorporation of the new cells. Fluoxetine increased BDNF levels in the hippocampus of both sexes. BDNF levels correlated with cell proliferation in males but not females. Brain and plasma levels indicated that females metabolized fluoxetine faster than males and produced more of the metabolite norfluoxetine. These data suggest that fluoxetine acts on multiple areas of the brain to increase cell proliferation, and the pattern of activation differs between males and females. Sex-specific effects of fluoxetine on neurotrophin mobilization and pharmacokinetics may contribute to these differences in neural plasticity.