Biology subjects

Herman, M. A.

Publications and source records attributed to Herman, M. A..

18 recordsLinked to original sources

bioRxiv2026

Manganese availability determines insulin sensitivity by enhancing Akt activity

Insulin signaling is a critical determinant of metabolic health, and impairments in insulin action contribute to the development of type 2 diabetes. The kinase Akt is a central mediator of insulin signaling and is required for insulin's suppression of hepatic glucose output. Although the regulation of Akt by the insulin receptor-PI3K pathway is well understood, there are instances in which signaling downstream of Akt is dissociated from proximal insulin signaling, for example in insulin resistance. Nonetheless, little is known about PI3K-independent mechanisms of Akt regulation. Here, we discovered that hepatocyte manganese concentrations are a key determinant of PI3K-independent Akt function in vivo. We further demonstrated that manganese increases Akt's catalytic efficiency, and quantitative phosphoproteomics revealed that manganese and insulin act additively to enhance Akt activity. Moreover, we uncovered that hepatic manganese concentrations fluctuate during fasting and feeding via carbohydrate-dependent transcriptional regulation of the manganese efflux transporter Slc30a10. This dynamic metal-signaling axis provides a mechanistic link between nutrient status and Akt activation, and suggests a molecular explanation for the glucose-lowering effects of manganese observed in humans. Our findings establish manganese as a physiologically regulated cofactor for Akt and position metal bioavailability as a previously unrecognized layer of insulin signaling control.

physiology↗

bioRxiv2025

A water-based platform test of paired or group social dominance and neural activity in male and female mice

Social hierarchy is an evolutionarily-conserved phenomenon determined by social dominance behavior that has profound influence on health and relevance in neuropsychiatric disorders. Despite this, the neural mechanisms underlying social dominance remain unclear and current behavioral tests are limited. Here, we describe a novel platform test of social dominance where mice compete for space on a small, elevated platform surrounded by cold water and rank is calculated by total time spent on platform. To validate this assay, we conducted tube test followed by platform test in cages of male and female mice. We observed stable rank in both sexes using the platform test and saw notable differences in overall cage hierarchy between tube and platform. Motivated behavior as measured by attempts to get on the platform was reduced over time. Cooperative behavior as measured by shared time on platform was observed across all ranks but predominantly demonstrated by subordinate mice. Corticosterone levels were significantly higher in females but showed no rank-specific differences. Neuronal activity in the prefrontal cortex was not rank-dependent, however activity in the habenula scaled with rank in both sexes and was significantly correlated with dominance in males. We also investigated group social dynamics using the platform test and identified stable hierarchies with more exaggerated dominance compared to paired platform testing. These results introduce the platform test as a novel method for assessing social dominance behavior in male and female mice, which can be used to further our understanding of the neurobiology underlying social hierarchy.

neuroscience↗

bioRxiv2025

TARP γ-8 is a target of ethanol that regulates self-administration and relapse in mice

BackgroundBehavioral pathologies that characterize alcohol use disorder (AUD) are driven by the powerful reinforcing, or rewarding, properties of the drug. We have shown that glutamate AMPA receptor (AMPAR) activity is both necessary and sufficient for alcohol (ethanol) reinforcement. Transmembrane AMPAR regulatory protein (TARP) {gamma}-8 is an essential auxiliary protein that regulates AMPAR expression and activity; however, the role of TARP {gamma}-8 in AUD or other forms of addiction remains largely unexplored. ObjectivesThis study investigated the mechanistic role of TARP {gamma}-8 in operant ethanol self-administration (model of primary reinforcement) and cue-induced reinstatement of ethanol-seeking behavior (model of conditioned reinforcement) using TARP {gamma}-8 heterozygous null (+/-) mice. To determine if TARP {gamma}-8 signaling is targeted by ethanol use, we evaluated protein expression of TARP {gamma}-8, GluA1, CaMKII, and PSD-95 following ethanol self-administration. ResultsA battery of tests evaluating food and water intake, taste reactivity, anxiety-like behavior, and object recognition memory showed no fundamental behavioral deficits in TARP {gamma}-8 (+/-) mice, and no differences in response to acute ethanol or home-cage drinking as compared to wild-types. However, TARP {gamma}-8 (+/-) mice exhibited significantly reduced acquisition and escalation of operant ethanol self-administration and reduced cue-induced reinstatement of ethanol-seeking behavior, with no differences in parallel sucrose-only controls. In wild-type mice, ethanol self-administration increased TARP {gamma}-8 expression in the amygdala, nucleus accumbens, and hippocampus, and increased GluA1 expression in the amygdala and prefrontal cortex, compared to sucrose controls. ConclusionThese findings highlight the specificity of TARP {gamma}-8 regulation of ethanol reinforcement mechanisms and identify this crucial AMPAR auxiliary protein as a target of ethanol in reward-related brain regions, highlighting its potential for development of novel pharmacotherapies for AUD.

neuroscience↗

bioRxiv2024

Glycerol-3-phosphate activates ChREBP, FGF21 transcription and lipogenesis in Citrin Deficiency

Citrin Deficiency (CD) is caused by inactivation of SLC25A13, a mitochondrial membrane protein required to move electrons from cytosolic NADH to the mitochondrial matrix in hepatocytes. People with CD do not like sweets. We discovered that SLC25A13 loss causes accumulation of glycerol-3-phosphate (G3P), which activates carbohydrate response element binding protein (ChREBP) to transcribe FGF21, which acts in the brain to restrain intake of sweets and alcohol, and to transcribe key genes of de novo lipogenesis. Mouse and human data establish G3P-ChREBP as a new mechanistic component of the Randle Cycle that contributes to metabolic dysfunction-associated steatotic liver disease (MASLD) and forms part of a system that communicates metabolic states from liver to brain in a manner that alters food and alcohol choices. The data provide a framework for understanding FGF21 induction in varied conditions, suggest ways to develop FGF21-inducing drugs, and drug candidates for both lean MASLD and support of urea cycle function in CD.

molecular biology↗

bioRxiv2024

Integration of metabolomic and transcriptomic analyses reveals novel regulatory functions of the ChREBP transcription factor in energy metabolism

Carbohydrate Response Element-Binding Protein (ChREBP) is a transcription factor that activates key genes involved in glucose, fructose, and lipid metabolism in response to carbohydrate feeding, but its other potential roles in metabolic homeostasis have not been as well studied. We used liver-selective GalNAc-siRNA technology to suppress expression of ChREBP in rats fed a high fat/high sucrose diet and characterized hepatic and systemic responses by integrating transcriptomic and metabolomic analyses. GalNAc-siChREBP-treated rats had lower levels of multiple short-chain acyl CoA metabolites compared to rats treated with GalNAc-siCtrl containing a non-targeting siRNA sequence. These changes were related to a sharp decrease in free CoA levels in GalNAc-siChREBP treated-rats, accompanied by lower expression of transcripts encoding enzymes and transporters involved in CoA biosynthesis. These activities of ChREBP likely contribute to its complex effects on hepatic lipid and energy metabolism. While core enzymes of fatty acid (FA) oxidation are induced by ChREBP knockdown, accumulation of liver acylcarnitines and circulating ketones indicate diversion of acetyl CoA to ketone production rather than complete oxidation in the TCA cycle. Despite strong suppression of pyruvate kinase and activation of pyruvate dehydrogenase, pyruvate levels were maintained, likely via increased expression of pyruvate transporters, and decreased expression of lactate dehydrogenase and alanine transaminase. GalNAc-siChREBP treatment increased hepatic citrate and isocitrate levels while decreasing levels of distal TCA cycle intermediates. The drop in free CoA levels, needed for the 2-ketoglutarate dehydrogenase reaction, as well as a decrease in transcripts encoding the anaplerotic enzymes pyruvate carboxylase, glutamate dehydrogenase, and aspartate transaminase likely contributed to these effects. GalNAc-siChREBP treatment caused striking increases in PRPP and ZMP/AICAR levels, and decreases in GMP, IMP, AMP, NaNM, NAD(P), and NAD(P)H levels, accompanied by reduced expression of enzymes that catalyze late steps in purine and NAD synthesis. ChREBP suppression also increased expression of a set of plasma membrane amino acid transporters, possibly as an attempt to replenish TCA cycle intermediates. In sum, combining transcriptomic and metabolomic analyses has revealed regulatory functions of ChREBP that go well beyond its canonical roles in control of carbohydrate and lipid metabolism to now include mitochondrial metabolism and cellular energy balance.

physiology↗

bioRxiv2024

Single residues in the complexin N-terminus exhibit distinct phenotypes in synaptic vesicle fusion

The release of neurotransmitters at central synapses is dependent on a cascade of protein interactions, specific to the presynaptic compartment. Amongst those dedicated molecules the cytosolic complexins play an incompletely defined role as synaptic transmission regulators. Complexins are multidomain SNARE complex binding proteins which confer both inhibitory and stimulatory functions. Using systematic mutagenesis and combining reconstituted in vitro membrane fusion assays with electrophysiology in neurons, we deciphered the function of the N-terminus of complexin II (Cpx). The N-terminus (amino acid 1 - 27) starts with a region enriched in hydrophobic amino acids (1-12), which can lead to lipid binding. In contrast to mutants which maintain the hydrophobic character and the stimulatory function of Cpx, non-conservative exchanges largely perturbed spontaneous and evoked exocytosis. Mutants in the downstream region (amino acid 11-18) show differential effects. Cpx-A12W increased spontaneous release without affecting evoked release whereas replacing D15 with amino acids of different shapes or hydrophobic properties (but not charge) not only increased spontaneous release, but also impaired evoked release and surprisingly reduced the size of the readily releasable pool, a novel Cpx function, unanticipated from previous studies. Thus, the exact amino acid composition of the Cpx N-terminus fine tunes the degree of spontaneous and evoked neurotransmitter release. Significance StatementWe describe in this work the importance of the N-terminal domain of the small regulatory cytosolic protein complexin in spontaneous and evoked glutamatergic neurotransmitter release at hippocampal mouse neurons. We show using a combination of biochemical, imaging and electrophysiological techniques that the binding of the proximal region of complexin (amino acids 1-10) to lipids is crucial for spontaneous synaptic vesicular release. Furthermore, we identify a single amino acid at position D15 which is structurally important since it not only is involved in spontaneous release but, when mutated, also decreases drastically the readily releasable pool, a function that was never attributed to complexin.

neuroscience↗

bioRxiv2023

Metformin Impairs Intestinal Fructose Metabolism

ObjectiveTo investigate the effects of metformin on intestinal carbohydrate metabolism in vivo. Method: Male mice preconditioned with a high-fat, high-sucrose diet were treated orally with metformin or a control solution for two weeks. Fructose metabolism, glucose production from fructose, and production of other fructose-derived metabolites were assessed using stably labeled fructose as a tracer. ResultsMetformin treatment decreased intestinal glucose levels and reduced incorporation of fructose-derived metabolites into glucose. This was associated with decreased intestinal fructose metabolism as indicated by decreased enterocyte F1P levels and diminished labeling of fructose-derived metabolites. Metformin also reduced fructose delivery to the liver. Proteomic analysis revealed that metformin coordinately down-regulated proteins involved carbohydrate metabolism including those involved in fructolysis and glucose production within intestinal tissue. ConclusionMetformin reduces intestinal fructose metabolism, and this is associated with broad-based changes in intestinal enzyme and protein levels involved in sugar metabolism indicating that metformins effects on sugar metabolism are pleiotropic. HighlightsO_LIMetformin decreases intestinal fructose absorption, metabolism, and fructose delivery to the liver. C_LIO_LIMetformin reduces intestinal glucose production from fructose-derived metabolites. C_LIO_LIMetformin reduces protein levels of multiple metabolic enzymes involved in fructose and glucose metabolism in intestinal tissue. C_LI

physiology↗

bioRxiv2022

Psychedelic compounds directly excite 5-HT2A Layer 5 Pyramidal Neurons in the Prefrontal Cortex through a 5-HT2A Gq-mediated activation mechanism

Psilocin, the active compound in Psilocybe sp. mushrooms, is a serotonergic psychedelic that has recently gained renewed interest due to its potential as a therapeutic tool. Despite promising clinical findings, the underlying signaling mechanisms and brain region-specific effects of psilocin and other psychedelic drugs remain unclear. Psilocin, like other psychedelic compounds, is an agonist at many serotonin and other biogenic amine receptors; however, activation of serotonin (5-Hydroxytryptamine, or 5-HT) 2A receptors (5-HT2ARs) is understood as the main molecular target for the psychoactive effects in animals and humans. 5-HT2ARs are abundantly expressed in the prefrontal cortex (PFC); however, the biochemical actions of psilocin on PFC neurons remain poorly understood. In this study, we used in vitro slice electrophysiology to examine how psilocin acutely alters the activity and electrophysiological properties of layer 5 pyramidal neurons in the mouse PFC. Focal application of psilocin (10M) onto nonspecified Layer 5 Pyramidal neurons in the prelimbic PFC of C57BL/6J mice produced variable effects on firing (increase, decrease, or no change). 5-HT2AR layer 5 pyramidal neurons in the mouse prelimbic PFC were identified via labeling in a 5-HT2A-ERT2-Cre mouse crossed with an Ai9 tdTomato reporter. Focal application of psilocin increased firing in all identified 5-HT2AR neurons but did not result in any significant changes in synaptic transmission. Overall, the results demonstrate that psilocin evokes strong firing changes in the PFC that are 5-HT2AR and Gq dependent, thereby providing valuable insights into the effects of psilocin on a brain region implicated in mediating psychedelic drug actions.

neuroscience↗

bioRxiv2022

Sex-Specific Changes in Neuronal Activity and Affective Behavior after Chronic Alcohol Consumption in CRF1-cre Transgenic Rats

Alcohol use disorders (AUDs) are characterized by compulsive alcohol use, loss of control over intake, and a negative emotional state during abstinence. While AUDs are associated with both mood and chronic pain disorders, the relationship between these associations remains unclear. Corticotropin releasing factor-1 receptor (CRF1) has been implicated in alcohol (EtOH) use, affective states, and pain sensitivity; often in a sex-dependent manner. Using CRF1-cre transgenic rats, we found no sex differences in basal affective behavior with the exception of mechanical sensitivity, where females were more sensitive to mechanical stimuli. Following baseline testing, rats began EtOH (or water) drinking under intermittent access conditions. Females consumed more alcohol in the first week, but overall EtOH intake was not significantly different between males and females. Following 3-4 weeks of drinking, rats were tested again for negative affect. EtOH drinking decreased mechanical sensitivity, but no other group effects were observed. However, individual EtOH intake was directly correlated with anxiety- and depressive-like behavior in both sexes. Interestingly, EtOH intake inversely correlated with thermal sensitivity in males only. There were no group differences in CRF1+ neuronal activity in either prelimbic or infralimbic cortices, but final session EtOH intake was significantly correlated with activity in CRF1+ neurons in the infralimbic cortex. Together, our results suggest complex interplay between affective state, EtOH drinking, and the role of prefrontal cortex CRF1-containing neurons in mediating these behaviors. Additionally, these results highlight the importance of examining individual differences in AUD-related behaviors. SIGNIFICANCE STATEMENTDespite alcohol use disorders being extremely comorbid with mood and pain disorders, there is still a limited understanding of the interaction and directionality between the them. To investigate this problem, rats were tested for affective behavior before and after being allowed to drink alcohol for 6 weeks. While baseline behavior did not predict subsequent intake, alcohol intake predicted both anxiety- and depressive-like behavior. These findings were accompanied by increased activity of the corticotropin releasing factor 1 containing neurons in the infralimbic region of the prefrontal cortex. Together, these findings reveal a new mechanism for understanding alcohol use.

neuroscience↗

bioRxiv2022

Sex differences in the impact of electronic nicotine vapor on corticotropin-releasing factor receptor 1 neurons in the mouse ventral tegmental area

Nicotine engages dopamine neurons in the ventral tegmental area (VTA) to encode reward and drive the development of nicotine addiction, however how nicotine selectively alters other VTA populations remains to be determined. Here, we used male and female CRF1-GFP mice and nicotine vapor exposure to examine the effects of nicotine in VTA corticotropin-releasing factor receptor 1 (CRF1) neurons. We use immunohistochemistry and electrophysiology to examine neuronal activity, excitability, and inhibitory signaling. We found that VTA CRF1 neurons are mainly dopaminergic and project to the nucleus accumbens (VTA-NAcCRF1 neurons). VTA-NAcCRF1 neurons show greater phasic inhibition in naive females and greater focal nicotine-induced increases in firing in naive males. Following acute nicotine vapor exposure, phasic inhibition was not altered, but focal nicotine-induced tonic inhibition was enhanced in females and diminished in males. Acute nicotine vapor exposure did not affect firing in VTA-NAcCRF1 neurons, but females showed lower baseline firing and higher focal nicotine-induced firing. Activity (cFos) was increased in the CRF1 dopaminergic VTA population in both sexes, but with greater increases in females. Following chronic nicotine vapor exposure, both sexes displayed reduced basal phasic inhibition and the sex difference in tonic inhibition following acute vapor exposure was no longer observed. Additionally, activity of the CRF1 dopaminergic VTA population was no longer elevated in either sex. These findings reveal sex- and exposure-dependent changes in mesolimbic VTA-NAc CRF1 neuronal activity, inhibitory signaling, and nicotine sensitivity following nicotine vapor exposure. These changes potentially contribute to nicotine-dependent behaviors and the intersection between stress, anxiety, and addiction.

neuroscience↗

bioRxiv2022

Sex Differences in Central Amygdala Reactivity following Psychedelic Drug Exposure

Psilocybin, and its active metabolite psilocin, have been shown to elicit rapid and long-lasting symptom improvements in a variety of affective psychiatric illnesses. However, the region-specific alterations underlying these therapeutic effects remain relatively unknown. The central amygdala (CeA) is a primary output region within the extended amygdala that is dysregulated in affective psychiatric disorders. Here, we measured CeA activity using the activity marker c-Fos and CeA reactivity using fiber photometry paired with an aversive air-puff stimulus. We found that psilocin administration acutely increased CeA activity in both males and females and increased stimulus specific CeA reactivity in females, but not males. In contrast, psilocin produced time-dependent decreases in reactivity in males, but not females as early as 2-days and lasting to 28-days post administration. We also measured behavioral responses to the air-puff stimulus and found sex-dependent changes in threat responding but not exploratory behavior or general locomotion. Repeated presentations of the auditory component of the air-puff were also performed and sex-specific effects of psilocin on CeA reactivity to the auditory-alone stimulus were also observed. This study provides new evidence that a single dose of psilocin produces sex-specific, time-dependent, and enduring changes in CeA reactivity and behavioral responding to specific components of an aversive stimulus.

neuroscience↗

bioRxiv2022

Paradoxical activation of SREBP1c and de novo lipogenesis by hepatocyte-selective ACLY depletion in obese mice.

Hepatic steatosis associated with high fat diets, obesity and type 2 diabetes is thought to be the major driver of severe liver inflammation, fibrosis, and cirrhosis. Cytosolic acetyl-coenzyme A (AcCoA), a central metabolite and substrate for de novo lipogenesis (DNL), is produced from citrate by ATP-citrate lyase (ACLY) and from acetate through AcCoA synthase short chain family member 2 (ACSS2). However, the relative contributions of these two enzymes to hepatic AcCoA pools and DNL rates in response to high fat feeding is unknown. We report here that hepatocyte-selective depletion of either ACSS2 or ACLY caused similar 50% decreases in liver AcCoA levels in obese mice, showing that both pathways contribute to generation of this DNL substrate. Unexpectedly however, the hepatocyte ACLY depletion in obese mice paradoxically increased total DNL flux measured by D2O incorporation into palmitate, while in contrast ACSS2 depletion had no effect. The increase in liver DNL upon ACLY depletion was associated with increased expression of nuclear sterol regulatory element-binding protein 1c (SREBP1c) and of its target DNL enzymes. This upregulated DNL enzyme expression explains the increased rate of palmitate synthesis in ACLY depleted livers. Furthermore, this increased flux through DNL may also contribute to the observed depletion of AcCoA levels due to its increased conversion to Malonyl CoA (MalCoA) and palmitate. Together, these data indicate that in HFD fed obese mice, hepatic DNL is not limited by its immediate substrates AcCoA or MalCoA, but rather by activities of DNL enzymes.

molecular biology↗

bioRxiv2022

The Intestine is a Major Contributor to Circulating TCA Cycle Intermediates in Mice

The tricarboxylic acid (TCA) cycle is the epicenter of cellular aerobic metabolism. TCA cycle intermediates facilitate energy production and provide anabolic precursors, but also function as intra- and extracellular metabolic signals regulating pleiotropic biological processes. Despite the importance of circulating TCA cycle metabolites as signaling molecules, the source of circulating TCA cycle intermediates remains uncertain. We observe that in mice, the concentration of TCA cycle intermediates in the portal blood exceeds that in tail blood indicating that the gut is a major contributor to circulating TCA cycle metabolites. With a focus on succinate as a representative of TCA cycle intermediate with signaling activities and using a combination of germ-free mice and isotopomer tracing, we demonstrate that intestinal microbiota are not major contributors to circulating TCA cycle metabolites. Moreover, we demonstrate that the endogenous succinate production is markedly higher than intestinal succinate absorption in normal physiological conditions. Altogether, these results indicate that endogenous succinate production within the intestinal tissue is a major physiological source of circulating succinate. These results provide a foundation for investigation into the role of intestine in regulating circulating TCA cycle metabolites and related signaling effects in health and disease.

physiology↗

bioRxiv2021

Sex Differences in Affective States and Association with Voluntary Ethanol Intake in Sprague Dawley Rats

Alcohol use disorders (AUDs) are a major problem across the United States. While AUD remains a complex human condition, it is difficult to isolate the directionality of anxiety and ethanol (EtOH) drinking from outside influences. The present study sought to investigate the relationship between affective states and EtOH intake using male and female Sprague Dawley rats. Using complementary tests of anxiety- and depressive-like behavior, we found sex- and test-specific differences in basal affective behavior such that females displayed enhanced anxiety-like behavior in the Splash Test and males displayed enhanced anxiety-like behavior in the Novelty Suppressed Feeding Test. Although there were no sex differences in EtOH intake and no correlation between anxiety-like behavior and subsequent EtOH intake, we did find that depressive-like behavior predicted future EtOH intake in females rats only. In addition, we observed an increase in depressive-like behavior is male rats in both the water and EtOH drinking groups. Furthermore, anxiety-like behavior, but not depressive-like behavior predicted subsequent EtOH intake in female rats. Lastly, we found a history of EtOH intake decreased pain thresholds in male and female rats. Together, these experiments provide important information on the complex interaction between negative affect and alcohol intake and how these two contexts reciprocally do, or do not, influence each other in a sex-specific manner.

neuroscience↗

bioRxiv2021

HGFAC is a ChREBP Regulated Hepatokine that Enhances Glucose and Lipid Homeostasis

Carbohydrate Responsive Element-Binding Protein (ChREBP) is a carbohydrate sensing transcription factor that regulates both adaptive and maladaptive genomic responses in coordination of systemic fuel homeostasis. Genetic variants in the ChREBP locus associate with diverse metabolic traits in humans, including circulating lipids. To identify novel ChREBP-regulated hepatokines that contribute to its systemic metabolic effects, we integrated ChREBP ChIP-seq analysis in mouse liver with human genetic and genomic data for lipid traits and identified Hepatocyte Growth Factor Activator (HGFAC) as a promising ChREBP-regulated candidate in mice and humans. HGFAC is a protease that activates the pleiotropic hormone Hepatocyte Growth Factor (HGF). We demonstrate that HGFAC KO mice have phenotypes concordant with putative loss-of-function variants in human HGFAC. Moreover, in gain- and loss-of-function genetic mouse models, we demonstrate that HGFAC enhances lipid and glucose homeostasis, in part, through actions to activate hepatic PPAR{gamma} activity. Together, our studies show that ChREBP mediates an adaptive response to overnutrition via activation of an HGFAC-HGF-PPAR{gamma} signaling axis in the liver to preserve glucose and lipid homeostasis.

physiology↗

bioRxiv2021

Sex- and Subtype-Specific Adaptations in Excitatory Signaling Onto Deep-Layer Prelimbic Cortical Pyramidal Neurons After Chronic Alcohol Exposure

Long-term alcohol use results in behavioral deficits including impaired working memory, elevated anxiety, and blunted inhibitory control that are associated with prefrontal cortical (PFC) dysfunction. Preclinical observations demonstrate multiple impairments in GABAergic neurotransmission onto deep-layer principal cells (PCs) in prelimbic cortex that suggest dependence-related cortical dysfunction is the product of elevated excitability in these cells. Despite accumulating evidence showing alcohol-induced changes in interneuron signaling onto PCs differ between sexes, there is limited data explicitly evaluating sex-specific ethanol effects on excitatory signaling onto deep-layer PCs that may further contribute to deficits in PFC-dependent behaviors. To address this, we conducted electrophysiological and behavioral tests in both male and female Sprague-Dawley rats to evaluate the effects of chronic ethanol exposure. Among our observations, we report a marked enhancement in glutamatergic signaling onto deep-layer PCs in male, but not female, rats after alcohol exposure. This phenomenon was furthermore specific to a sub-class of PC, sub-cortically projecting Type-A cells, and coincided with enhanced anxiety-like behavior, but no observable deficit in working memory. In contrast, female rats displayed an alcohol-induced facilitation in working memory performance with no change in expression of anxiety-like behavior. Together, these results suggest fundamental differences in alcohol effects on cell activity, cortical sub-circuits, and PFC-dependent behaviors across male and female rats.

neuroscience↗

bioRxiv2021

Cellular, physiological, and behavioral validation of a CRFR1:Cre-tdTomato transgenic rat for use in basic neuroscience research

Corticotropin-releasing factor type-1 (CRF1) receptors are critical to stress responses because they allow neurons to respond to CRF released in response to stress. Our understanding of the precise role of CRF1-expressing neuronal populations in CRF-mediated behaviors has been largely limited to mouse experiments due to the lack of genetic tools available to selectively visualize and manipulate CRF1+ cells in rats. Here, we describe the generation and validation of a transgenic CRF1:Cre-tdTomato rat, which expresses a bicistronic iCre-2A-tdTomato transgene directed by 200kb of promoter and enhacer sequence surrounding the Crhr1 cDNA present within a BAC clone, that has been transgenically inserted into the rat genome. We report that Crhr1 and Cre mRNA expression are highly colocalized in CRF1:Cre-tdTomato rats within both the central amygdala (CeA), composed of mostly GABAergic neurons, and in the basolateral amygdala (BLA), composed of mostly glutamatergic neurons. In the CeA, membrane properties, inhibitory synaptic transmission, and responses to CRF bath application in tdTomato+ neurons are similar to those previously reported in GFP+ cells in CRFR1-GFP mice. We show that stimulatory DREADD receptors can be selectively targeted to CeA CRF1+ cells via virally delivered Cre-dependent transgenes, that transfected Cre/tdTomato+ cells are activated by clozapine-n-oxide in vitro and in vivo, and that activation of these cells in vivo increases anxiety-like behavior and nocifensive responses. Given the accuracy of expression in the CRF1:Cre rat, modern genetic techniques used to investigate the anatomy, physiology, and behavioral function of CRF1+ neurons and circuits can now be performed in assays that require the use of rats as the model organism. Impact StatementA novel transgenic rat for studying the role of specific corticotropin-releasing factor type-1 receptor-expressing cell populations in physiology and behavior.

neuroscience↗

bioRxiv2020

Chemogenetic stimulation of mouse central amygdala corticotropin-releasing factor neurons: Effects on cellular and behavioral correlates of alcoholdependence

BackgroundCorticotropin-releasing factor (CRF) signaling in the central nucleus of the amygdala (CeA) plays a critical role in rodent models of excessive alcohol drinking. However, the source of CRF acting in the CeA during alcohol withdrawal remains to be identified. In the present study, we hypothesized that CeA CRF interneurons may represent a behaviorally relevant source of CRF to the CeA increasing motivation for alcohol via negative reinforcement. MethodsWe tested this hypothesis in male mice and used chemogenetics to stimulate CeA CRF neurons in vitro and in vivo. ResultsWe first observed that Crh mRNA expression in the anterior part of the mouse CeA, at the junction with the interstitial nucleus of the posterior limb of the anterior commissure, correlates positively with alcohol intake in C57BL/6J males with a history of chronic binge drinking. We then found that chemogenetic activation of CeA CRF neurons in Crh-IRES-Cre mouse brain slices increases gamma-aminobutyric acid (GABA) release in the medial CeA in part via CRF1 receptor activation, indicating local CRF release. While chemogenetic stimulation of CeA CRF neurons exacerbated novelty-induced feeding suppression, as seen in C57BL/6J males withdrawn from chronic intermittent alcohol inhalation, it had no effect on voluntary alcohol consumption, following either acute or chronic manipulation. ConclusionsAltogether, these findings indicate that hyperactivity of CeA CRF neurons may contribute to elevated CeA GABA levels and negative affect during alcohol withdrawal but is not sufficient to drive alcohol intake escalation in dependent mice.

neuroscience↗