Corticostriatal mechanisms of action learning and habit formation

National Institute On Alcohol Abuse And Alcoholism

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Abstract

Gestational Ethanol Alters Behavioral Flexibility and Striatal Cholinergic Transmission Exposure to EtOH during gestation has undesirable effects on many neural functions, including attention and cognition. These EtOH effects, generally known as alcohol related neurodevelopmental disorders (ARND) are one of the major toxin-related causes of developmental disabilities in humans. The levels of EtOH exposure that produce ARND are not yet known, and factors such as timing and pattern of exposure alter the likelihood of neural impairment. Our previous studies demonstrated that EtOH exposure throughout pregnancy and the early postnatal period in mice (gestational ethanol exposure, GEE) resulted in loss of stimulus-response (e.g. habit) learning, as well as hyperactive instrumental behavior (but not general locomotor hyperactivity). These cognitive/behavioral impairments appeared to involve reduced GABAergic synaptic transmission and altered endocannabinoid signaling in dorsal striatum. The firing pattern of dorsal striatal MSNs related to stimulus-response learning were also disrupted following GEE. These findings indicate roles for disrupted DS function in altered cognition/operant behavior induced by early life alcohol exposure. We further explored GEE effects on striatum and related behaviors, with an emphasis on cognitive flexibility. In these experiments we focused on EtOH exposure during the early postnatal period (P0-P10) in mice, as brain development during this period is roughly equivalent to the last trimester in humans. We used inhalational EtOH exposure to pups with their mothers as in our past studies. The blood alcohol levels in pups averaged 150 mg/dl, well above intoxicating levels in adult mice. Control mice were exposed to air in the same chambers. We noted that the time for mothers to retrieve pups was slightly slower in the GEE than in the air-exposed control group. Furthermore, the weight of female GEE mice was lower at P60 compared to the air-exposed group, but male weights were not different across groups. These findings indicate that early postnatal EtOH exposure has differential effects on the development of female and male mice, perhaps due to deficits in maternal care. We next tested adult mice from the GEE and Air groups in an instrumental lever-pressing learning-reversal-extinction task with two levers present in the chamber. The mice first learned to press one of two levers on a self-paced fixed ratio 1 (FR1) schedule. They were then trained on an FR5 schedule. No between-group differences were observed in learning and lever-pressing rates in either sex under either FR schedule. Mice were then trained on a reversal task in which presses on the originally non-rewarded lever were now rewarded, while pressing the original rewarded lever did not result in reward. Both female and male GEE mice showed reversal performance similar to air controls. We next examined lever-pressing during extinction (i.e. no reward for pressing either lever). Male GEE mice showed performance comparable to air controls. However, female GEE mice exhibited greater lever pressing on the first extinction day compared to air controls. This finding indicates that female mice may persist in performing previously rewarded actions. To determine if skill learning and performance that involve the striatum is altered following GEE, we trained adult mice in the accelerating rotarod task. A 4-40 RPM acceleration was used and mice were trained for 5 trials per day over 4 days. Female and male mice from both groups showed improved performance over the first two training days. However, female mice in the GEE group showed poorer performance on days 3 and 4 compared to air controls. The pattern of these deficits was interesting, as the female GEE mice showed faster falling times on the apparatus in the first trials on days 3 and 4 compared to the last trials on the previous days. In contrast the air control group females showed little to no impairment on these trials relative to their previous day late trial performance. To determine if GEE mice were losing skill expertise over time we instituted a one week delay after training day 4 and then tested the mice again on a "retention" day consisting of 4 trials. The air control females showed a slight loss in performance on trial 1 of retention with near full recovery by trial 2. The GEE females, on the other hand, exhibited severe impairment (i.e. inability to stay on the apparatus) on the first retention trial and did not fully recover performance until trial 3-4. The rotarod performance was not correlated with animal weight. Male mice showed no treatment-related differences in rotarod performance on any trial or day, and indeed their performance was optimal on the later trials each day. These findings indicate that female mice exposed to EtOH early in development are deficient in retaining skill performance and can only achieve control-level performance with additional training. These findings may have implications for deficits in procedural learning in ARND, particularly in females. Given the role of striatal neurotransmitters in rotarod learning and performance, we have begun examining effects of GEE on cholinergic, dopaminergic and glutamatergic transmission in dorsolateral striatum of mice in the different treatment groups. To measure dopamine release in striatal slices, we used fast scan cyclic voltammetry with electrical afferent stimulation. Similar to the behavioral experiments, we observed sex differences in the effect of GEE on dopamine release. Female GEE mice showed enhanced dopamine release compared to air controls, while male mice showed no treatment-related difference. Electrical stimulation-induced dopamine release in striatal slices involves activation of acetylcholine release from cholinergic interneurons that subsequently activates nicotinic acetylcholine receptors on dopaminergic terminals. To determine if a change in this cholinergic drive might contribute to the GEE-induced change in dopamine release we applied the nicotinic receptor antagonist Dihyro beta erythroidin (DHbeta) to slices and measured the resulting decrease in dopamine release. The inhibition of dopamine release by DHbetaE was larger in the GEE female slices in comparison to those from the air control group. The GEE-induced alteration in the role of acetylcholine in dopamine release indicated that we should examine alterations in the function of striatal cholinergic interneurons and acetylcholine release. We then measured GEE effects on excitability of striatal cholinergic using whole-cell electrophysiological recording in brain slices from female mice. We observed that GEE increased depolarization-induced action potential firing frequency in these neurons, potentially related to an increase in the hyperpolarization-induced cation current that regulates excitability of these neurons. We also measured electrical stimulation-induced acetylcholine release in female mice using brain slice photometry and the newly developed genetically-encoded fluorescent sensor called GRABACh3.0. These experiments revealed that GEE alters the duration of release in the presence of an inhibitor of the acetylcholine-catabolizing enzyme acetylcholine esterase. Overall, our findings indicate that GEE impairs instrumental extinction and skill learning in a sex-dependent manner, with prominent effects in female mice. Enhanced striatal dopamine release driven by a larger role for striatal acetylcholine may contribute to these behavioral deficits. We hope to examine how manipulations of this striatal circuitry might mimic or rescue these behavioral phenotypes. Some of these findings have been posted in a preprint by Bariselli et al. in bioRxiv.