Caspase-dependent ablation of indirect medium spiny neurons projecting to external globus pallidus promotes compulsive ethanol-seeking and drinking behaviors

Caspase-dependent ablation of indirect medium spiny neurons projecting to external globus pallidus promotes compulsive ethanol-seeking and drinking behaviors | bioRxiv /* */ /* */ <!-- <!-- /*! * yepnope1.5.4 * (c) WTFPL, GPLv2 */ (function(a,b,c){function d(a){return"[object Function]"==o.call(a)}function e(a){return"string"==typeof a}function f(){}function g(a){return!a||"loaded"==a||"complete"==a||"uninitialized"==a}function h(){var a=p.shift();q=1,a?a.t?m(function(){("c"==a.t?B.injectCss:B.injectJs)(a.s,0,a.a,a.x,a.e,1)},0):(a(),h()):q=0}function i(a,c,d,e,f,i,j){function k(b){if(!o&&g(l.readyState)&&(u.r=o=1,!q&&h(),l.onload=l.onreadystatechange=null,b)){"img"!=a&&m(function(){t.removeChild(l)},50);for(var d in y[c])y[c].hasOwnProperty(d)&&y[c][d].onload()}}var j=j||B.errorTimeout,l=b.createElement(a),o=0,r=0,u={t:d,s:c,e:f,a:i,x:j};1===y[c]&&(r=1,y[c]=[]),"object"==a?l.data=c:(l.src=c,l.type=a),l.width=l.height="0",l.onerror=l.onload=l.onreadystatechange=function(){k.call(this,r)},p.splice(e,0,u),"img"!=a&&(r||2===y[c]?(t.insertBefore(l,s?null:n),m(k,j)):y[c].push(l))}function j(a,b,c,d,f){return q=0,b=b||"j",e(a)?i("c"==b?v:u,a,b,this.i++,c,d,f):(p.splice(this.i++,0,a),1==p.length&&h()),this}function k(){var a=B;return a.loader={load:j,i:0},a}var l=b.documentElement,m=a.setTimeout,n=b.getElementsByTagName("script")[0],o={}.toString,p=[],q=0,r="MozAppearance"in l.style,s=r&&!!b.createRange().compareNode,t=s?l:n.parentNode,l=a.opera&&"[object Opera]"==o.call(a.opera),l=!!b.attachEvent&&!l,u=r?"object":l?"script":"img",v=l?"script":u,w=Array.isArray||function(a){return"[object Array]"==o.call(a)},x=[],y={},z={timeout:function(a,b){return b.length&&(a.timeout=b[0]),a}},A,B;B=function(a){function b(a){var a=a.split("!"),b=x.length,c=a.pop(),d=a.length,c={url:c,origUrl:c,prefixes:a},e,f,g;for(f=0;f<d;f++)g=a[f].split("="),(e=z[g.shift()])&&(c=e(c,g));for(f=0;f<b;f++)c=x[f](c);return c}function g(a,e,f,g,h){var i=b(a),j=i.autoCallback;i.url.split(".").pop().split("?").shift(),i.bypass||(e&&(e=d(e)?e:e[a]||e[g]||e[a.split("/").pop().split("?")[0]]),i.instead?i.instead(a,e,f,g,h):(y[i.url]?i.noexec=!0:y[i.url]=1,f.load(i.url,i.forceCSS||!i.forceJS&&"css"==i.url.split(".").pop().split("?").shift()?"c":c,i.noexec,i.attrs,i.timeout),(d(e)||d(j))&&f.load(function(){k(),e&&e(i.origUrl,h,g),j&&j(i.origUrl,h,g),y[i.url]=2})))}function h(a,b){function c(a,c){if(a){if(e(a))c||(j=function(){var a=[].slice.call(arguments);k.apply(this,a),l()}),g(a,j,b,0,h);else if(Object(a)===a)for(n in m=function(){var b=0,c;for(c in a)a.hasOwnProperty(c)&&b++;return b}(),a)a.hasOwnProperty(n)&&(!c&&!--m&&(d(j)?j=function(){var a=[].slice.call(arguments);k.apply(this,a),l()}:j[n]=function(a){return function(){var b=[].slice.call(arguments);a&&a.apply(this,b),l()}}(k[n])),g(a[n],j,b,n,h))}else!c&&l()}var h=!!a.test,i=a.load||a.both,j=a.callback||f,k=j,l=a.complete||f,m,n;c(h?a.yep:a.nope,!!i),i&&c(i)}var i,j,l=this.yepnope.loader;if(e(a))g(a,0,l,0);else if(w(a))for(i=0;i (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0];var j=d.createElement(s);var dl=l!='dataLayer'?'&l='+l:'';j.src='//www.googletagmanager.com/gtm.js?id='+i+dl;j.type='text/javascript';j.async=true;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-M677548'); Skip to main content Home About Submit ALERTS / RSS Search for this keyword Advanced Search New Results Caspase-dependent ablation of indirect medium spiny neurons projecting to external globus pallidus promotes compulsive ethanol-seeking and drinking behaviors Humza Haroon , Matthew Baker , View ORCID Profile Doo-Sup Choi doi: https://doi.org/10.1101/2025.02.21.639573 Humza Haroon 1 Department of Molecular Pharmacology and Experimental Therapeutics, Mayo Clinic College of Medicine and Science , Rochester, MN 55905, USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Matthew Baker 1 Department of Molecular Pharmacology and Experimental Therapeutics, Mayo Clinic College of Medicine and Science , Rochester, MN 55905, USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site Doo-Sup Choi 1 Department of Molecular Pharmacology and Experimental Therapeutics, Mayo Clinic College of Medicine and Science , Rochester, MN 55905, USA 2 Department of Psychiatry and Psychology, Mayo Clinic College of Medicine and Science , Rochester, MN 55905, USA 3 Neuroscience Program, Mayo Clinic College of Medicine and Science , Rochester, MN 55905, USA Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Doo-Sup Choi For correspondence: choids{at}mayo.edu Abstract Full Text Info/History Metrics Supplementary material Preview PDF Abstract The dorsomedial striatum (DMS) is primarily recognized for regulating goal-directed reward-seeking behaviors, while the dorsolateral striatum (DLS) is predominantly associated with movement and habitual behaviors. In this study, we sought to investigate two pathways, direct medium spiny neuron (dMSN) and indirect medium spiny neuron (iMSN) in the two dorsal striatal subregions (DMS and DLS) in ethanol-seeking and drinking behaviors. Here, we selectively ablated iMSN DMS→GPe and iMSN DLS→GPe and trained mice to exhibit goal-directed and habitual reward-seeking behaviors using random ratio (RR) and random interval (RI) operant conditioning, respectively. We found that partial ablation of iMSN DLS→GPe exhibited increased resilience to bitter-tasting ethanol solution when subjected to quinine adulterated reward in operant conditioning paradigms, suggesting compulsive-like seeking behavior. Consistently, in a separate cohort of mice, we found that the iMSN DLS→GPe ablated mice show higher preference and consumption of quinine adulterated ethanol solution than control mice in two-bottle choice continuous access drinking, with increasing quinine concentration and exhibit more compulsive-like behavior. On the other hand, ablation of iMSN DMS→GPe resulted in insensitivity to satiety-based reward devaluation in RR-trained mice, consistent with a shift toward habitual behavior but no change in compulsive ethanol-seeking and drinking behavior. Together, our findings demonstrate that DLS iMSN function is essential in inhibiting compulsive-like behavior. Introduction The basal ganglia are a group of subcortical nuclei, of which coordinated activity allows decision-making, reward-related learning and motor actions ( Schultz et al., 1997 ; Packard and Knowlton, 2002 ; Yin and Knowlton, 2006 ). In humans, the dorsal striatum (DS) is anatomically separated by the internal capsule forming the caudate and putamen nucleus whereas, in rodents, DS is a homogeneous structure with overlapping axonal connectivity due to an absence of anatomical border ( Hoover et al., 2003 ; Alloway et al., 2006 ; Alloway et al., 2009 ; Hooks et al., 2018 ). However, the rodent DS has two functionally recognizable subregions, the dorsomedial striatum (DMS, equivalent to caudate in primates and humans) and the dorsolateral striatum (DLS, equivalent to putamen) ( Hauber and Schmidt, 1994 ; Graybiel, 2008 ). The DMS plays an essential role in the acquisition and expression of goal-directed behavior controlled by action-outcome (A-O) contingency, while the DLS is primarily involved in habitual behaviors regulated by stimulus-response (S-R) association ( Voorn et al., 2004 ; Yin and Knowlton, 2006 ; Graybiel, 2008 ). The striatum consists of GABAergic medium spiny neurons (MSN), making up more than 95% of the neuronal population, divided into equal proportions of direct pathway MSN (dMSN) and indirect pathway MSN (iMSN) ( Gerfen and Surmeier, 2011 ). These two morphologically indistinguishable striatal MSN have extensive dendritic arbors providing synaptic integration from upstream neurons ( Kawaguchi et al., 1990 ). In the classic model of basal ganglia, direct and indirect pathways have been considered to play opposing roles, with the direct pathway sending the “go signal” for action promotion, while the indirect pathway sends the “no go signal” to inhibit or suppress the action ( Albin et al., 1989 ; Alexander and Crutcher, 1990 ; DeLong, 1990 ; Mink, 2018 ). In general, dMSN stimulation enhances reinforcement learning, whereas iMSN stimulation induces avoidance behaviors and inhibits reinforcement ( Lobo and Nestler, 2011 ; Kravitz et al., 2012 ). However, accumulating evidence suggests that both dMSN and iMSN are co-activated during movement and action selection ( Cui, 2013 ; Tecuapetla et al., 2014 ). This suggests that arbitration of the activities between the two MSNs determines the direction of ongoing actions. Consequently, a higher iMSN activation is associated with action termination ( Cui, 2013 ; Geddes et al., 2018 ). Although studies have investigated the distinct DMS vs. DLS or iMSN vs. dMSN functions, it remains unclear if iMSN may have specialized functions in inhibiting compulsive reward-seeking within each striatal subregion. Compulsive-like alcohol drinking refers to persistent alcohol drinking despite negative consequences, the hallmark of defining alcohol use disorder (AUD) in the Diagnostic and Statistical Manual of Mental Disorders 5 (DSM 5) ( Grant et al., 2015 ). This compulsive aspect is a major obstacle in treating AUD ( Dawson et al., 2005 ; Moos and Moos, 2006 ; Sinha, 2009 ; Koob and Volkow, 2010 ; Hopf and Lesscher, 2014 ). Thus, understanding how the activity of iMSN within each dorsal striatal region contributes to compulsive behavior may help guide the target and development of new therapeutic strategies. Here, we selectively ablate iMSN circuits in the DMS and DLS to investigate their roles in compulsive reward-seeking and drinking behaviors using escalating concentrations of quinine adulterated ethanol solution in operant conditioning and two-bottle choice drinking tasks. Our findings indicate that the DLS iMSN function is essential in inhibiting compulsive-like behavior. Methods Animals All experimental procedures were approved by the Mayo Clinic Institutional Animal Care and Use Committee and performed following National Institute of Health (NIH) guidelines. C57BL/6J male and female mice were purchased from Jackson Laboratory (Bar Harbor, ME). D1-tdT/D2-eGFP bitransgenic mice were bred in the lab. Mice were housed in standard Plexiglas cages. The colony room was maintained at a constant temperature (24 ± 1°C) and humidity (60 ± 2%) with a 12 hr light/dark cycle (lights on at 06:00 A.M.). We used 8- to 10-week-old mice for all experiments. Mice were allowed ad libitum access to food and water. For the operant conditioning tests, mice were food-restricted to 85% of their baseline weight, at which time they were maintained for the duration of experimental procedures. Operant conditioning We conducted operant conditioning using the same operant chambers/schedules as our previous studies ( Hong et al., 2019 ; Kang et al., 2020 ; Baker et al., 2023 ) (Supplemental Fig. S1B). Briefly, mice were placed in operant chambers (Med-Associates, St Albans, VT) in which they poked a single hole for an outcome of ethanol-containing solution (10 μl per reinforcement). For all experiments, sweetened ethanol (10% sucrose+10% ethanol, 10S10E) was used as the reward. Before training, mice were food-restricted to approximately 85% body weight, which was maintained for the duration of experimental procedures. On the first day, mice were trained to approach the reward magazine with a reward delivered on a random time schedule for 30 min. Next, mice were trained on a fixed ratio 1 schedule for 30 min in 7 sessions. After acquiring nose-poking behavior, mice were trained on random interval (RI10 1-2days/RI30 5 days/RI60 3 days) to develop habitual behavior or random ratio (RR2 1–2 days/RR5 5 days/RR10 3 days) to form goal-directed reward-seeking. Sessions were completed after 30 min or following 60 reward reinforcements. RI10/30/60 delivered one reward outcome on average every 10/30/60s after the last reward outcome to develop habitual reward-seeking. RR2/5/10 delivered one reward outcome on average every 2/5/10 response in the correct nose-poke hole to develop goal-directed reward-seeking. Reward devaluation and extinction test On the devalued day, mice were given 1 hour of ad libitum access to the outcome (10S10E) previously earned by nose poking (for devaluation) or food pellets and then underwent serial nonreinforced extinction sessions in each training context. The order of the valuation context was counterbalanced across mice and was 10 min in duration. According to the sensory-specific satiety theory, we conducted reward devaluation for 1 hr and the extinction test for 10 min to determine which is more dominant between goal-directed behavior and habit. We provided unlimited food chow to make the reward worthwhile for the valued state, whereas we provided an unlimited 10S10E solution to devaluate the outcome value. We conducted the extinction tests after the last sessions of RR10, and RI60. Quinine adulteration test Mice were placed in the RI60 operant conditioning task with increasing concentrations of quinine in the 10S10E solution (0, 100, 250 µM quinine). Mice performed two RI60 sessions at each quinine concentration and data was averaged between the two sessions. Stereotaxic surgery for virus injection Mice were anesthetized with isoflurane (3.5% in oxygen gas) using a VetFlo TM vaporizer with a single-channel anesthesia stand (Kent Scientific Corporation, Torrington, CT) and placed on the digital stereotaxic alignment system (model 1900; David Kopf instruments). Hair was trimmed, and the skull was exposed using 8-gauge electrosurgical skin cutter (KLS martin, Jacksonville, FL). The skull was leveled using a dual-tilt measurement tool. Holes were drilled in the skull at the appropriate stereotaxic coordinates. Viruses were infused to the DLS (AP + 0.7 mm, ML +/- 2.5 mm, DV –3.1 mm from bregma), DMS (AP + 0.7 mm, ML +/- 1.25 mm, DV –3.1 mm from bregma) or GPe (AP –0.4 mm, ML +/- 2.0 mm, DV −3.8 mm from bregma) at 100 nl/min for 4 minutes (400 nl total) through a 33-gauge injection needle (cat # NF33BV; World Precision Instruments) using a microsyringe pump (Model UMP3; World Precision Instruments). The injection needle remained in place for an additional 5 min following the end of the injection. All viruses were purchased from Addgene and were injected at the following titers: retrograde AAV-Ef1a-mCherry-IRES-Cre (7×10 12 vg/mL), AAV5-flex-taCasp3-TEVp (7×10 12 vg/mL), AAV-Ef1a-DIO-EYFP (7×10 12 vg/mL). Following stereotaxic surgery, we injected buprenorphine extended release (1 mg/kg, S.C.; ZooPharm, Laramie, WY, USA) to alleviate post-surgery pain. Caspase 3-mediated circuit ablation To ablate the circuit in a targeted fashion, we used a genetically engineered caspase 3 (Casp3), whose activation commits a cell to apoptosis ( Chelur and Chalfie, 2007 ). Endogenous caspase 3 exists as procaspase 3, which is cleaved into its active form by upstream apoptotic signals and other caspase proteins. This genetically engineered caspase lacks the cleavage site for upstream caspases and can only be cleaved by a tobacco etch virus protease (TEVp) which is coexpressed in an AAV ( Yang et al., 2013 ). AAV-flex-taCasp3-TEVp was expressed in a cre-dependent manner in the DLS to only ablate cells that express Cre recombinase from retrograde injections in the GPe (AAV-Ef1a-mCherry-IRES-Cre). Importantly, caspase 3 triggers cell-autonomous apoptosis, minimizing the risk of off-target effects and toxicity to neighboring cells ( Mallet et al., 2002 ; Smart et al., 2017 ; Patton et al., 2021 ). Immunofluorescence Brains were fixed with 4% paraformaldehyde (Sigma-Aldrich, St. Louis, MO) and transferred to 30% sucrose (Sigma-Aldrich) in phosphate-buffered saline at 4 °C for 72 hr. Brains were then frozen in dry ice and sectioned at 40 µm using a microtome (Leica Corp., Bannockburn, IL). Brain slices were stored at −20 °C in a cryoprotectant solution containing 30% sucrose (Sigma-Aldrich) and 30% ethylene glycol (Sigma-Aldrich) in phosphate-buffered saline. Sections were incubated in 0.2% Triton X-100 (Sigma-Aldrich), 5% bovine serum albumin in phosphate-buffered saline for 1 hr, followed by incubation with the primary antibody in 5% bovine serum albumin overnight at 4 °C. After three washes in phosphate-buffered saline, the sections were mounted onto a glass slide coated with gelatin and cover-slipped with a VECTASHIELD® antifade mounting medium (Vector Laboratories, Burlingame, CA). Images were obtained using an LSM 780 laser scanning confocal microscope (Carl Zeiss, Heidelberg, Germany) using a 10x or 40x water-immersion lens. Cell count values were determined using Image J (1.53t, National Institute of Health, USA). We used 509 (eGFP), 581 (tdTomato), 610 (mCherry), and 405 channels for imaging. Open field test The open-field test (OFT) was conducted in chambers (Med-Associates, St Albans, VT) to measure the locomotor responses of mice. The session lasted 30 minutes and total distance and velocities were recorded using beam breaks. Animals were habituated to the testing room in their home cage 1 hour prior to behavioral testing. Two-bottle choice continuous access ethanol-drinking Oral ethanol-containing solution (10S10E) consumption and preference were examined using a two-bottle choice test in the mouse home cage. Mice were individually housed and given 24 hr access to two bottles: water and 10S10E. Every other day, the bottles were measured, and bottle positions were switched to prevent place preference development. After baseline consumption and preference were established for 8 days, mice underwent two surgeries 1 week apart. After 3 weeks of the second injection, mice were single-housed in the home cage for the two-bottle choice test. Mice were given 24 hr access to two bottles: water and 10S10E but with increasing concentrations of quinine in the 10S10E solution (0, 100, 250 µM quinine). Mice performed two sessions (4 days, 2 days on each side) at each quinine concentration and data was averaged between the two sessions. 10S10E and water consumption were normalized for evaporation ( Nam et al., 2013 ; Leon et al., 2023 ). Briefly, the total volume of the liquid evaporated was calculated by averaging 2-days of evaporation from the four control cages without mice. Then, the volume of water or ethanol that evaporated was subtracted from the total consumption of water or ethanol for each mouse. Data analysis All data are represented as mean ± SEM using Prism 9.0 (GraphPad Software, San Diego, CA). The statistical significance was set at p < 0.05. The statistical significance was set at p < 0.05. All p values less than 0.05 were marked as “*” in the graphs. Detailed statistical tests and data with exact p values are listed in Supplemental Table S1. Results iMSNs primarily project to the GPe The dorsal striatal projections can be divided into two distinct pathways: direct pathway medium spiny neurons (dMSN) target the entopeduncular nucleus (EP; rodent homolog of the primate internal globus pallidus), and substantia nigra pars reticulata (SNr) directly, whereas indirect pathway medium spiny neurons (iMSN) project to the external part of the globus pallidus (GPe), which then send axons to the output nuclei via the subthalamic nucleus (STN) ( Alexander and Crutcher, 1990 ). To measure the effect of ablation, we injected retrograde mCherry virus into the GPe and counted the mCherry+ cells in the DLS and DMS of both control and casp3 ablated mice. We found that casp3 ablation indeed significantly reduces the neurons present within the DLS and DMS (Supplemental Fig. S2B and S2D, Unpaired t test, p < 0.05). Several studies have revealed that dMSN has axon collaterals in the GPe (Fujiyama et al., 2011; Kawaguchi et al., 1990 ; Wu et al., 2000). To this, we injected the retrograde mCherry virus into the GPe and counted iMSN in the DMS and DLS of both control and casp3 mice of bitransgenic D1-tdT/D2-eGFP mice. We found a significantly reduced number of iMSN in both DLS and DMS with a greater ablation of iMSN in the DLS ( Fig. 1B and 1D , Unpaired t test, p < 0.05). This suggests that iMSN from the dorsal striatum primarily project to the GPe and the effect on behavior we observe is due to the partial ablation of these iMSN. Download figure Open in new tab Figure 1. Effect of Caspase 3 (casp3)-dependent ablation of dorsal striatum iMSN. A , Schematic of virus injections for Cre-dependent ablation of DLS neurons and representative DLS images, dMSN (red) iMSN (green) of sham control and caspase mice (Scale: 50 µm) from n = 3 mice/group. B , Caspase mice showed a significant reduction of iMSN in the DLS (t = 3.87, p = 0.018). C , Schematic of virus injections for Cre-dependent ablation of DMS neurons and representative DMS images, dMSN (red) iMSN (green), of sham control and caspase mice (Scale: 50 µm) from n = 3 mice/group. D , Caspase mice showed a significant reduction of iMSN in the DMS (t = 3.32, p = 0.029). Figure ( A ) and ( C ) was created with BioRender.com . Caspase3-dependent ablation of iMSN DLS→GPe promotes increased compulsive-like ethanol-seeking but not motor suppression To determine whether ablation of the iMSN DLS→GPe circuit modulates goal-directed or habitual behavior, we used a Cre-dependent caspase 3 virus, which induces cell-autonomous death with minimal toxicity to neighboring cells (Yang; Mallet et al., 2002 ; Chelur and Chalfie, 2007 ; Smart et al., 2017 ; Patton et al., 2021 ). We bilaterally injected a mCherry-tagged retrograde virus expressing Cre recombinase [AAV-Ef1a-mCherry-IRES-Cre] into the GPe, followed by a second injection of Cre-dependent caspase-3 (AAV5-flex-taCasp3-TEVp; or AAV5-Ef1a-DIO EYFP (control) into the DLS (Supplemental Fig. S1A). We validated a significant reduction in mCherry-positive neurons in the DLS of the caspase group (unpaired t- test, p < 0.05; Supplemental Fig. S2B). Since the DLS has long been associated with motor function, we examined if DLS iMSN ablation resulted in motor dysfunction. In the open field test (Supplemental Fig. S3A), we observed no significant differences in spontaneous locomotion, velocity, or time spent in the center zone, indicating that DLS iMSN ablation does not alter basic motor function (Mann-Whitney U test, p > 0.05; Supplemental Fig. S3B). In the devaluation test, both control and caspase mice in the RR group showed significant reduction in nose pokes for the devalued state (Wilcoxon test, p < 0.05; Fig. 2B ), consistent with goal-directed behavior. In contrast, both control and caspase mice in the RI group showed no significant differences between the valued and devalued states (Wilcoxon test, p > 0.05; Fig. 2C ) indicating habitual behavior. Altogether our results demonstrate iMSN circuit ablation does not impair overall performance or acquisition rate in the operant reward-seeking task. Download figure Open in new tab Figure 2. Effect of Caspase 3 (casp3)-dependent ablation of dorsolateral striatum (DLS) indirect medium spiny neurons (iMSN) projecting to external globus pallidus (GPe) on goal-directed and habitual alcohol-seeking. A , Schematic of evaluation test to determine reward-outcome valuation testing. B , Control and Casp3 animals showed significantly decreased nosepoke behavior in the devalued state, indicating goal-directed behavior. C , Control and Casp3 animals showed no differences between the valued and devalued states, indicating habitual behavior. D , Casp3 mice showed increased compulsive ethanol-seeking behaviors in quinine adulterated ethanol-seeking tasks. Wilcoxon test ( B-C ). n = 8 −12/group. Data represent mean ± SEM. Two-way repeated measures ANOVA with Tukey’s posthoc tests were used for ( D ). See Table S1 for full statistical information. Figure ( A ) was created with BioRender.com . Next, we examined whether the iMSN DLS→GPe ablation influenced compulsive-like seeking behavior. In the quinine adulteration test, all mice displayed reductions in reward-seeking behaviors as the quinine concentration increased (RM Two-way ANOVA, p < 0.05; Fig. 2D ), showing sensitivity to the aversive taste of quinine. However, caspase mice showed increased magazine entries compared to control mice in both RR and RI groups, indicating more compulsive-like seeking behaviors (RM Two-way ANOVA, p < 0.05; Fig. 2D ). Collectively, this indicates that decreased iMSN DLS→GPe activity contributes to the resilience to bitter tasting quinine containing ethanol solution, suggestive of compulsive-like ethanol seeking behaviors. Caspase3-dependent ablation of iMSN DMS→GPe shifts mice toward habitual-behavior To determine whether ablation of iMSN DMS→GPe modulates goal-directed or habitual reward-seeking, we injected a mCherry-tagged retrograde Cre-virus into the GPe, followed by a second injection of Cre-dependent caspase-3 (or control virus) into the DMS (Supplemental Fig. S2C). We saw a significant reduction in mCherry-positive neurons in the DMS in the caspase group (unpaired t test, p < 0.05; Supplemental Fig. S3D). Previous studies have implicated iMSN in the DMS being associated with motor function ( Kravitz et al., 2012 ; Cui et al., 2021 ), so we sought to determine if partial iMSN ablation resulted in motor dysfunction. In the open field test (Supplemental Fig. 3A), we observed no significant differences in spontaneous locomotion, velocity, or time spent in the center zone, indicating that DMS iMSN ablation does not alter basic motor function (Mann-Whitney test, p > 0.05; Supplemental Fig. S3C). In the devaluation test, control mice in the RR group showed a reduction in extinction session nose pokes for the devalued state (Wilcoxon test, p < 0.05; Fig. 3B ), consistent with goal-directed behavior. Consistent but conversely to our previous findings where activation of iMSN in DMS shifted habitual to goal-directed behavior (Kang; Kang et al., 2020 ), we find interestingly, RR caspase mice exhibit habitual behavior during devalued state ( p > 0.05; Fig. 3B ), indicating that loss of iMSN DLS→GPe circuit function promotes a shift from goal-directed to habitual behavior. In contrast, RI caspase mice showed no significant differences between the valued and devalued states in both control and caspase mice ( p > 0.05; Fig. 3C ). Overall, this indicates that DMS iMSN ablation specifically reduces goal-directed behavior without changing habitual behaviors during conditioned reward. Download figure Open in new tab Figure 3. Effect of Caspase 3 (casp3)-dependent ablation of dorsolateral striatum (DLS) indirect medium spiny neurons (iMSN) projecting to external globus pallidus (GPe) on goal-directed and habitual alcohol-seeking. A , Schematic of evaluation test to determine reward-outcome valuation testing. B , Control mice showed significantly decreased nose poke behavior in the devalued state, indicating goal-directed behavior. However, Casp3 mice showed a shift to habitual behavior. C , Control and Casp3 animals showed no differences between the valued and devalued states, indicating habitual behaviors. D , Control and Casp3 mice showed no difference in compulsive ethanol-seeking behaviors in quinine adulterated ethanol-seeking tasks. Wilcoxon test ( B-C ). n = 8/group. Data represent mean ± SEM. Two-way repeated measures ANOVA with Tukey’s posthoc tests were used for ( D ). See Table S1 for full statistical information. Figure ( A ) was created with BioRender.com . Caspase3-dependent ablation of iMSN DLS → GPe promotes compulsive ethanol-drinking in mice Recurrent use of ethanol despite negative consequences is a hallmark feature of compulsive alcohol drinking in AUD. Given that we found increased compulsive-like alcohol-seeking in iMSN DLS→GPe ablated mice, we asked whether iMSN ablation would increase voluntary ethanol intake and preference. We first established baseline ethanol intake which showed distinct sex-specific drinking behaviors. While both, male and female, mice show no difference in preference for ethanol, female mice exhibit higher consumption of ethanol (Supplemental Fig. S4). We then examined the effect of partial ablation of iMSN from the two dorsal striatal regions. To this, we subjected 10 mice (5 male + 5 female) per group to the two-bottle choice continuous access ethanol drinking test. We found that increasing concentrations of quinine reduced ethanol consumption and preference for both caspase and control groups showing sensitivity to bitter taste of quinine. Consistent with our previous findings, we found that iMSN DLS→GPe ablated mice showed higher preference than the control group (RM Two-way ANOVA, Fig. 4B ). While we did not see a group difference in consumption between the caspase and control group for the iMSN DLS→GPe ablated mice, we did see a significantly higher consumption in the caspase group at 250 µM concentration of quinine, again showing resilience to the bitter tasting ethanol solution exhibiting more compulsive-like behavior. The iMSN DMS→GPe control and caspase group showed no group differences in ethanol preference or consumption (RM Two-way ANOVA, Fig. 4C ). Together, these results suggest that reduced iMSN DLS→GPe activity contributes to compulsive-like ethanol drinking. Download figure Open in new tab Figure 4. Effect of Caspase 3 (casp3)-dependent ablation of dorsal striatum indirect medium spiny neurons (iMSN) projecting to external globus pallidus (GPe) on ethanol-drinking. A , Two-bottle choice continuous access paradigm showing the increasing concentration of quinine in 10S10E. B , Partial ablation of iMSN DLS→GPe (Casp3 mice) promotes compulsive ethanol-drinking behaviors in C57BL/6J mice. C, Partial ablation of iMSN DMS→GPe (Casp3 mice) showed no change in ethanol-drinking behaviors in C57BL/6J mice. n = 8/group. Data represent mean ± SEM. Two-way repeated measures ANOVA with Tukey’s posthoc tests were used for ( B , C ). See Table S1 for full statistical information. Figure ( A ) was created with BioRender.com . Discussion In this study, we demonstrate that DLS iMSN plays a critical role in suppressing compulsive-like behaviors. Specifically, we observed that partial ablation of iMSN DLS→GPe promotes compulsive ethanol-seeking and drinking behaviors in C57BL/6J mice. Based on other studies ( Lobo and Nestler, 2011 ; Kravitz et al., 2012 ), we hypothesized that DLS iMSN dysfunction would further promote reward-seeking behaviors. However, our results indicated that reward devaluation was not sensitive enough to show any changes in reward-seeking following circuit ablation. Previously, we showed that activation of astrocytes in the DMS increases iMSN activity, which leads to a shift from habitual to goal-directed behavior ( Kang et al., 2020 ). Conversely, partial iMSN ablation in the DMS leads to a shift from goal to habit in mice trained in the random ratio (RR) paradigm ( Figure 3B ). Previously, studies in humans have shown that hyperactive striatum is associated with compulsive behavior ( Saxena and Rauch, 2000 ; Menzies et al., 2008 ). Similarly, preclinical studies using various mouse models have also revealed that a hyperactive striatum is involved in compulsive-like behavior ( Welch et al., 2007 ; Ahmari et al., 2013 ; Burguiere et al., 2013 ). Hyperactive direct pathway or a reduction in activation of indirect pathway have long been suggested to be the underlying mechanisms of compulsive behaviors ( Graybiel and Rauch, 2000 ; Saxena and Rauch, 2000 ; Maia et al., 2008 ). Additionally, an imbalance between the two pathways has been shown to be associated with compulsive drug-seeking ( Lobo and Nestler, 2011 ; Bock et al., 2013 ; Roltsch Hellard et al., 2019 ). Consistent with previous studies, our study presents the notion that reduced indirect pathway activity increases compulsive-like seeking and drinking behaviors. The role of iMSN in the DLS are critical for regulating compulsive-like seeking and drinking behaviors. The ablation of these neurons disrupts the balance between the direct and indirect pathways, leading to the disinhibition of reward-driven actions. This imbalance manifests as increased compulsive-like behaviors, including excessive ethanol seeking and drinking. Interestingly, we found a significant increase in magazine entries in the quinine adulteration task while there were no differences in acquisition of goal-directed or habitual behavior ( Figure 2 ). Furthermore, DLS iMSN ablation led to disinhibition of the bitter reward and continued to show significantly higher ethanol consumption during the two-bottle choice continuous access ( Figure 4 ). This suggests that segregating possible effects on reward-seeking and reward-consumption may be critical to understanding unique effects in different behavioral models of habitual or compulsive behavior ( Pegg et al., 2021 ; Wright and Wesson, 2021 ). The ability to change behavior in response to signals from the environment and to stop previously effective actions is known as behavioral flexibility ( Kim and Hikosaka, 2015 ; Uddin, 2021 ). It facilitates problem-solving and enables modification of behavior to change the environment. According to Griffin and Guez ( Griffin and Guez, 2014 ), behavioral flexibility includes a variety of skills, such as the capacity to perform an existing behavior in a novel setting, block a previously rewarded activity, and create new behavior ( Griffin and Guez, 2014 ; Kim and Hikosaka, 2015 ; Uddin, 2021 ). Since behavioral flexibility is the ability to stop or control unnecessary or risky actions, proper activation of iMSN is required for goal-directed behaviors and voluntary movements. Overall, our findings support striatal iMSN role in reward aversion and compulsive-like behavior. In conclusion, our results suggest that iMSN inputs are a critical mechanism for controlling the expression of compulsive behaviors toward alcohol use. The partial ablation of iMSN in the dorsal striatum leads to increased compulsive-like seeking and drinking behavior by reducing inhibitory control, which in turn leads to reduced behavior flexibility. Conflict of Interest All the authors declare that the research was conducted without any commercial or financial relationships that could be construed as a potential conflict of interest. Author Contributions Humza Haroon: Conceptualization, Investigation, Formal analysis, Writing – Original Draft, Writing – Review & Editing, Visualization. Matthew Baker: Conceptualization, Writing – Review & Editing, Visualization. Doo-Sup Choi: Resources, Writing – Review & Editing, Supervision, Project administration, Funding acquisition. Acknowledgment We thank all the laboratory members for their helpful discussion and comments. Figures were created with BioRender.com . This research was supported by the Samuel C. Johnson for Genomics of Addiction Program at Mayo Clinic, the Ulm Foundation, and the National Institute of Health (AA029258, AG072898). Footnotes Conflict of Interest: The authors declare no competing financial interests. References ↵ Ahmari SE , Spellman T , Douglass NL , Kheirbek MA , Simpson HB , Deisseroth K , Gordon JA , Hen R ( 2013 ) Repeated cortico-striatal stimulation generates persistent OCD-like behavior . Science 340 : 1234 – 1239 . OpenUrl Abstract / FREE Full Text ↵ Albin RL , Young AB , Penney JB ( 1989 ) The functional anatomy of basal ganglia disorders . Trends Neurosci 12 : 366 – 375 . OpenUrl CrossRef PubMed Web of Science ↵ Alexander GE , Crutcher MD ( 1990 ) Functional architecture of basal ganglia circuits: neural substrates of parallel processing . Trends Neurosci 13 : 266 – 271 . OpenUrl CrossRef PubMed Web of Science ↵ Alloway KD , Lou L , Nwabueze-Ogbo F , Chakrabarti S ( 2006 ) Topography of cortical projections to the dorsolateral neostriatum in rats: multiple overlapping sensorimotor pathways . J Comp Neurol 499 : 33 – 48 . OpenUrl CrossRef PubMed Web of Science ↵ Alloway KD , Smith JB , Beauchemin KJ , Olson ML ( 2009 ) Bilateral projections from rat MI whisker cortex to the neostriatum, thalamus, and claustrum: forebrain circuits for modulating whisking behavior . J Comp Neurol 515 : 548 – 564 . OpenUrl CrossRef PubMed Web of Science ↵ Baker M , Kang S , Hong SI , Song M , Yang MA , Peyton L , Essa H , Lee SW , Choi DS ( 2023 ) External globus pallidus input to the dorsal striatum regulates habitual seeking behavior in male mice . Nat Commun 14 : 4085 . OpenUrl CrossRef PubMed ↵ Bock R , Shin JH , Kaplan AR , Dobi A , Markey E , Kramer PF , Gremel CM , Christensen CH , Adrover MF , Alvarez VA ( 2013 ) Strengthening the accumbal indirect pathway promotes resilience to compulsive cocaine use . Nat Neurosci 16 : 632 – 638 . OpenUrl CrossRef PubMed ↵ Burguiere E , Monteiro P , Feng G , Graybiel AM ( 2013 ) Optogenetic stimulation of lateral orbitofronto-striatal pathway suppresses compulsive behaviors . Science 340 : 1243 – 1246 . OpenUrl Abstract / FREE Full Text ↵ Chelur DS , Chalfie M ( 2007 ) Targeted cell killing by reconstituted caspases . Proc Natl Acad Sci U S A 104 : 2283 – 2288 . OpenUrl Abstract / FREE Full Text ↵ Cui G ( 2013 ) Concurrent activation of striatal direct and indirect pathways during action initiation . Nature 494 : 238 – 242 . OpenUrl CrossRef PubMed Web of Science ↵ Cui Q , Pamukcu A , Cherian S , Chang IYM , Berceau BL , Xenias HS , Higgs MH , Rajamanickam S , Chen Y , Du X , Zhang Y , McMorrow H , Abecassis ZA , Boca SM , Justice NJ , Wilson CJ , Chan CS ( 2021 ) Dissociable Roles of Pallidal Neuron Subtypes in Regulating Motor Patterns . J Neurosci 41 : 4036 – 4059 . OpenUrl Abstract / FREE Full Text ↵ Dawson DA , Grant BF , Li TK ( 2005 ) Quantifying the risks associated with exceeding recommended drinking limits . Alcohol Clin Exp Res 29 : 902 – 908 . OpenUrl CrossRef PubMed Web of Science ↵ DeLong MR ( 1990 ) Primate models of movement disorders of basal ganglia origin . Trends Neurosci 13 : 281 – 285 . OpenUrl CrossRef PubMed Web of Science ↵ Geddes CE , Li H , Jin X ( 2018 ) Optogenetic editing reveals the hierarchical organization of learned action sequences . Cell 174 : 32 – 43 . OpenUrl CrossRef PubMed ↵ Gerfen CR , Surmeier DJ ( 2011 ) Modulation of striatal projection systems by dopamine . Annu Rev Neurosci 34 : 441 – 466 . OpenUrl CrossRef PubMed Web of Science ↵ Grant BF , Goldstein RB , Saha TD , Chou SP , Jung J , Zhang H , Pickering RP , Ruan WJ , Smith SM , Huang B , Hasin DS ( 2015 ) Epidemiology of DSM-5 Alcohol Use Disorder: Results From the National Epidemiologic Survey on Alcohol and Related Conditions III . JAMA Psychiatry 72 : 757 – 766 . OpenUrl CrossRef PubMed ↵ Graybiel AM ( 2008 ) Habits, rituals, and the evaluative brain . Annu Rev Neurosci 31 : 359 – 387 . OpenUrl CrossRef PubMed Web of Science ↵ Graybiel AM , Rauch SL ( 2000 ) Toward a neurobiology of obsessive-compulsive disorder . Neuron 28 : 343 – 347 . OpenUrl CrossRef PubMed Web of Science ↵ Griffin AS , Guez D ( 2014 ) Innovation and problem solving: a review of common mechanisms . Behav Processes 109 Pt B : 121 – 134 . OpenUrl CrossRef PubMed ↵ Hauber W , Schmidt WJ ( 1994 ) Differential effects of lesions of the dorsomedial and dorsolateral caudate-putamen on reaction time performance in rats . Behav Brain Res 60 : 211 – 215 . OpenUrl CrossRef PubMed ↵ Hong SI , Peyton L , Chern Y , Choi DS ( 2019 ) Novel Adenosine Analog, N6-(4-Hydroxybenzyl)-Adenosine, Dampens Alcohol Drinking and Seeking Behaviors . J Pharmacol Exp Ther 371 : 260 – 267 . OpenUrl Abstract / FREE Full Text ↵ Hooks BM , Papale AE , Paletzki RF , Feroze MW , Eastwood BS , Couey JJ , Winnubst J , Chandrashekar J , Gerfen CR ( 2018 ) Topographic precision in sensory and motor corticostriatal projections varies across cell type and cortical area . Nat Commun 9 : 3549 . OpenUrl CrossRef PubMed ↵ Hoover JE , Hoffer ZS , Alloway KD ( 2003 ) Projections from primary somatosensory cortex to the neostriatum: the role of somatotopic continuity in corticostriatal convergence . J Neurophysiol 89 : 1576 – 1587 . OpenUrl CrossRef PubMed Web of Science ↵ Hopf FW , Lesscher HM ( 2014 ) Rodent models for compulsive alcohol intake . Alcohol 48 : 253 – 264 . OpenUrl CrossRef PubMed Kang S Activation of astrocytes in the dorsomedial striatum facilitates transition from habitual to goal-directed reward-seeking behavior . ↵ Kang S , Hong SI , Lee J , Peyton L , Baker M , Choi S , Kim H , Chang SY , Choi DS ( 2020 ) Activation of Astrocytes in the Dorsomedial Striatum Facilitates Transition From Habitual to Goal-Directed Reward-Seeking Behavior . Biol Psychiatry 88 : 797 – 808 . OpenUrl CrossRef PubMed ↵ Kawaguchi Y , Wilson CJ , Emson PC ( 1990 ) Projection subtypes of rat neostriatal matrix cells revealed by intracellular injection of biocytin . J Neurosci 10 . ↵ Kim HF , Hikosaka O ( 2015 ) Parallel basal ganglia circuits for voluntary and automatic behaviour to reach rewards . Brain 138 : 1776 – 1800 . OpenUrl CrossRef PubMed ↵ Koob GF , Volkow ND ( 2010 ) Neurocircuitry of addiction . Neuropsychopharmacology 35 : 217 – 238 . OpenUrl CrossRef PubMed Web of Science ↵ Kravitz AV , Tye LD , Kreitzer AC ( 2012 ) Distinct roles for direct and indirect pathway striatal neurons in reinforcement . Nat Neurosci 15 : 816 – 818 . OpenUrl CrossRef PubMed ↵ Leon BE , Peyton L , Essa H , Wieden T , Marion N , Childers WE , Abou-Gharbia M , Choi DS ( 2023 ) A novel monobactam lacking antimicrobial activity, MC-100093, reduces sex-specific ethanol preference and depressive-like behaviors in mice . Neuropharmacology 232 : 109515 . OpenUrl CrossRef PubMed ↵ Lobo MK , Nestler EJ ( 2011 ) The striatal balancing act in drug addiction: distinct roles of direct and indirect pathway medium spiny neurons . Front Neuroanat 5 : 41 . OpenUrl CrossRef PubMed ↵ Maia TV , Cooney RE , Peterson BS ( 2008 ) The neural bases of obsessive-compulsive disorder in children and adults . Dev Psychopathol 20 : 1251 – 1283 . OpenUrl CrossRef PubMed Web of Science ↵ Mallet VO , Mitchell C , Guidotti JE , Jaffray P , Fabre M , Spencer D , Arnoult D , Kahn A , Gilgenkrantz H ( 2002 ) Conditional cell ablation by tight control of caspase-3 dimerization in transgenic mice . Nat Biotechnol 20 : 1234 – 1239 . OpenUrl CrossRef PubMed Web of Science ↵ Menzies L , Chamberlain SR , Laird AR , Thelen SM , Sahakian BJ , Bullmore ET ( 2008 ) Integrating evidence from neuroimaging and neuropsychological studies of obsessive-compulsive disorder: the orbitofronto-striatal model revisited . Neurosci Biobehav Rev 32 : 525 – 549 . OpenUrl CrossRef PubMed Web of Science ↵ Mink JW ( 2018 ) Basal ganglia mechanisms in action selection, plasticity, and dystonia . Eur J Paedriatr Neurol 22 : 225 – 229 . OpenUrl CrossRef ↵ Moos RH , Moos BS ( 2006 ) Rates and predictors of relapse after natural and treated remission from alcohol use disorders . Addiction 101 : 212 – 222 . OpenUrl CrossRef PubMed Web of Science ↵ Nam HW , Hinton DJ , Kang NY , Kim T , Lee MR , Oliveros A , Adams C , Ruby CL , Choi DS ( 2013 ) Adenosine transporter ENT1 regulates the acquisition of goal-directed behavior and ethanol drinking through A2A receptor in the dorsomedial striatum . J Neurosci 33 : 4329 – 4338 . OpenUrl Abstract / FREE Full Text ↵ Packard MG , Knowlton BJ ( 2002 ) Learning and memory functions of the Basal Ganglia . Annu Rev Neurosci 25 : 563 – 593 . OpenUrl CrossRef PubMed Web of Science ↵ Patton MS , Heckman M , Kim C , Mu C , Mathur BN ( 2021 ) Compulsive alcohol consumption is regulated by dorsal striatum fast-spiking interneurons . Neuropsychopharmacology 46 : 351 – 359 . OpenUrl CrossRef PubMed ↵ Pegg S , Jeong HJ , Foti D , Kujawa A ( 2021 ) Differentiating stages of reward responsiveness: Neurophysiological measures and associations with facets of the behavioral activation system . Psychophysiology 58 : e13764 . OpenUrl CrossRef PubMed ↵ Roltsch Hellard E , Binette A , Zhuang X , Lu J , Ma T , Jones B , Williams E , Jayavelu S , Wang J ( 2019 ) Optogenetic control of alcohol-seeking behavior via the dorsomedial striatal circuit . Neuropharmacology 155 : 89 – 97 . OpenUrl CrossRef PubMed ↵ Saxena S , Rauch SL ( 2000 ) Functional neuroimaging and the neuroanatomy of obsessive-compulsive disorder . Psychiatr Clin North Am 23 : 563 – 586 . OpenUrl CrossRef PubMed Web of Science ↵ Schultz W , Dayan P , Montague PR ( 1997 ) A neural substrate of prediction and reward . Science 275 : 1593 – 1599 . OpenUrl Abstract / FREE Full Text ↵ Sinha R ( 2009 ) Modeling stress and drug craving in the laboratory: implications for addiction treatment development . Addict Biol 14 : 84 – 98 . OpenUrl CrossRef PubMed Web of Science ↵ Smart AD , Pache RA , Thomsen ND , Kortemme T , Davis GW , Wells JA ( 2017 ) Engineering a light-activated caspase-3 for precise ablation of neurons in vivo . Proc Natl Acad Sci U S A 114 : E8174 – E8183 . OpenUrl Abstract / FREE Full Text ↵ Tecuapetla F , Matias S , Dugue GP , Mainen ZF , Costa RM ( 2014 ) Balanced activity in basal ganglia projection pathways is critical for contraversive movements . Nat Commun 5 : 4315 . OpenUrl CrossRef PubMed ↵ Uddin LQ ( 2021 ) Cognitive and behavioural flexibility: neural mechanisms and clinical considerations . Nat Rev Neurosci 22 : 167 – 179 . OpenUrl CrossRef PubMed ↵ Voorn P , Vanderschuren LJ , Groenewegen HJ , Robbins TW , Pennartz CM ( 2004 ) Putting a spin on the dorsal-ventral divide of the striatum . Trends Neurosci 27 : 468 – 474 . OpenUrl CrossRef PubMed Web of Science ↵ Welch JM , Lu J , Rodriguiz RM , Trotta NC , Peca J , Ding JD , Feliciano C , Chen M , Adams JP , Luo J , Dudek SM , Weinberg RJ , Calakos N , Wetsel WC , Feng G ( 2007 ) Cortico-striatal synaptic defects and OCD-like behaviours in Sapap3-mutant mice . Nature 448 : 894 – 900 . OpenUrl CrossRef PubMed Web of Science ↵ Wright KN , Wesson DW ( 2021 ) The tubular striatum and nucleus accumbens distinctly represent reward-taking and reward-seeking . J Neurophysiol 125 : 166 – 183 . OpenUrl CrossRef PubMed Yang CF Sexually dimorphic neurons in the ventromedial hypothalamus govern mating in both sexes and aggression in males . ↵ Yang CF , Chiang MC , Gray DC , Prabhakaran M , Alvarado M , Juntti SA , Unger EK , Wells JA , Shah NM ( 2013 ) Sexually dimorphic neurons in the ventromedial hypothalamus govern mating in both sexes and aggression in males . Cell 153 : 896 – 909 . OpenUrl CrossRef PubMed ↵ Yin HH , Knowlton BJ ( 2006 ) The role of the basal ganglia in habit formation . Nat Rev Neurosci 7 : 464 – 476 . OpenUrl CrossRef PubMed Web of Science View the discussion thread. Back to top Previous Next Posted February 25, 2025. Download PDF Supplementary Material Email Thank you for your interest in spreading the word about bioRxiv. NOTE: Your email address is requested solely to identify you as the sender of this article. Your Email * Your Name * Send To * Enter multiple addresses on separate lines or separate them with commas. You are going to email the following Caspase-dependent ablation of indirect medium spiny neurons projecting to external globus pallidus promotes compulsive ethanol-seeking and drinking behaviors Message Subject (Your Name) has forwarded a page to you from bioRxiv Message Body (Your Name) thought you would like to see this page from the bioRxiv website. Your Personal Message CAPTCHA This question is for testing whether or not you are a human visitor and to prevent automated spam submissions. Share Caspase-dependent ablation of indirect medium spiny neurons projecting to external globus pallidus promotes compulsive ethanol-seeking and drinking behaviors Humza Haroon , Matthew Baker , Doo-Sup Choi bioRxiv 2025.02.21.639573; doi: https://doi.org/10.1101/2025.02.21.639573 Share This Article: Copy Citation Tools Caspase-dependent ablation of indirect medium spiny neurons projecting to external globus pallidus promotes compulsive ethanol-seeking and drinking behaviors Humza Haroon , Matthew Baker , Doo-Sup Choi bioRxiv 2025.02.21.639573; doi: https://doi.org/10.1101/2025.02.21.639573 Citation Manager Formats BibTeX Bookends EasyBib EndNote (tagged) EndNote 8 (xml) Medlars Mendeley Papers RefWorks Tagged Ref Manager RIS Zotero Tweet Widget Facebook Like Google Plus One Subject Area Neuroscience Subject Areas All Articles Animal Behavior and Cognition (7624) Biochemistry (17651) Bioengineering (13873) Bioinformatics (41887) Biophysics (21424) Cancer Biology (18566) Cell Biology (25465) Clinical Trials (138) Developmental Biology (13365) Ecology (19871) Epidemiology (2067) Evolutionary Biology (24293) Genetics (15591) Genomics (22478) Immunology (17715) Microbiology (40331) Molecular Biology (17150) Neuroscience (88492) Paleontology (666) Pathology (2828) Pharmacology and Toxicology (4817) Physiology (7635) Plant Biology (15114) Scientific Communication and Education (2044) Synthetic Biology (4286) Systems Biology (9817) Zoology (2268)