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Abstract

The question of how the brain processes and records experiences for future recall is crucial to understanding learning and behavior. Traditionally, this has been thought to rely on synaptic plasticity – changes in the connections between individual neurons – that lead to the emergence of groups of cells that encode a particular salient feature known as the engram. Attention has recently been drawn to the fact that intrinsic plasticity, or experience-driven changes in the membrane excitability of individual neurons may also play a critical role in the proper function of learning. In light of this information, it is important to understand the mechanisms that contribute to this phenomenon. Activity of small-conductance calcium activated potassium (SK) channels, specifically their downregulation, is known to be required for the regulation of certain forms of cellular excitability such as firing rate and the size of the mAHP. In the cortex and hippocampus, activation of muscarinic acetylcholine receptors (mAChRs) has been shown both to inhibit SK channel activity, and facilitate both cellular excitability in vitro and acquisition of learned behavior in vivo. And yet, the precise mechanism by which SK channels are downregulated in response to activity during learning, and how this process is facilitated by mAChR activity remains unclear. In order to address these questions, experiments were performed using a combination of whole-cell patch-clamp recordings from Purkinje cells with pharmacological and genetic manipulations, as well as following a cerebellum-dependent associative learning task. Following tetanization with both a previously characterized somatic depolarization (SD) protocol, as well as a more physiologically relevant synaptic protocol, a stable, significant increase in the evoked firing rate, as well as a decrease in the SK2-channel mediated AHP minimum amplitude were observed. These effects were significantly enhanced with bath-application of a metabotropic agonist in conjunction with the synaptic protocol. Additionally, changes in cellular excitability were absent in recordings made using mice expressing a Purkinje-cell specific SK2 channel knockout. One downstream effect of Gq-signaling pathway is an increase in PKA activity, which is necessary for internalization of SK2 channels. Expression of intrinsic plasticity in Purkinje cells was also blocked in the presence of an intracellular PKA inhibitor. These experiments suggest one mechanism by which experience-dependent activity might lead to changes in the intrinsic excitability of neurons. To further characterize the role of intrinsic plasticity in encoding learning, recordings were made from mice two days after delay-eyeblink conditioning training. In recordings from animals that successfully acquired the conditioned response, a significant reduction in AHP was observed in response to both synaptic stimulus and evoked activity. Additionally, induction of intrinsic plasticity using the SD protocol was blocked in conditioned animals. Taken together the results of these experiments provide further insights into the the processes that lead to the induction and expression of intrinsic plasticity throughout the cerebellum. They also suggest that activity-driven changes in cellular excitability are not limited to a timescale of minutes or hours, but instead last on the order of days, and may therefore be relevant to help maintain learning.

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