Control of Ca²⁺flux between the cytosol and intracellular Ca²⁺stores is essential for maintaining normal cellular function. It has been well established in both neuronal and non-neuronal cells that stromal interaction molecule 1 (STIM1) initiates and regulates refilling Ca²⁺into the ER. Here, we describe a novel, additional role for STIM1, the regulation of free cytosolic Ca²⁺, and the consequent control of spike firing in neurons. Among central neurons, cerebellar Purkinje neurons express the highest level of STIM1, and they fire continuously in the absence of stimulation, making somatic Ca²⁺homeostasis of particular importance. By using Purkinje neuron-specific STIM1 knock-out (STIM1^PKO) male mice, we found that the deletion of STIM1 delayed clearance of cytosolic Ca²⁺in the soma during ongoing neuronal firing. Deletion of STIM1 also reduced the Purkinje neuronal excitability and impaired intrinsic plasticity without affecting long-term synaptic plasticity. In vestibulo-ocular reflex learning, STIM1^PKOmale mice showed severe deficits in memory consolidation, whereas they were normal in memory acquisition. Our results suggest that STIM1 is critically involved in the regulation of the neuronal excitability and the intrinsic plasticity of the Purkinje neurons as well as cerebellar memory consolidation.SIGNIFICANCE STATEMENTStromal interaction molecule 1 (STIM1), which regulates the refilling of ER Ca²⁺, has been investigated in several systems including the CNS. In addition to a previous study showing that STIM1 regulates dendritic ER Ca²⁺refilling and mGluR1-mediated synaptic transmission, we provide compelling evidence describing a novel role of STIM1 in spike firing Purkinje neurons. We found that STIM1 regulates cytosolic Ca²⁺clearance of the soma during spike firing, and the interruption of this cytosolic Ca²⁺clearing disrupts neuronal excitability and cerebellar memory consolidation. Our results provide new insights into neuronal functions of STIM1 from single neuronal Ca²⁺dynamics to behavior level.

Intrinsic plasticity of cerebellar Purkinje cells (PCs) has recently been demonstrated in cerebellar local circuits; however, its physiological impact on cerebellar learning and memory remains elusive. Here, we suggest that intrinsic plasticity of PCs is tightly involved in motor memory consolidation based on findings from PC-specific STIM1 knockout male mice, which show severe memory consolidation deficiency in vestibulo-ocular reflex memory. Gain-up training of the vestibulo-ocular reflex produced a decrease in the synaptic weight of PCs in both the WT and KO groups. However, intrinsic plasticity was impaired only in the knockout mice. Furthermore, the observed defects in the intrinsic plasticity of PCs led to the formation of aberrant neural plasticity in the vestibular nucleus neurons. Our results suggest that synergistic modulation of intrinsic and synaptic plasticity in PCs is required for the changes in downstream plasticity in the vestibular nucleus, and thereby contributing to the long-term storage of motor memory.SIGNIFICANCE STATEMENTSynaptic plasticity is a well-known mechanism for learning and memory. Although plasticity of excitability, intrinsic plasticity, of the cerebellar Purkinje cell has been reported in both directions (potentiation and depression), the physiological role of intrinsic plasticity still remains ambiguous. In this study, we suggest that both synaptic and intrinsic plasticity are required for successful memory consolidation in cerebellar eye movement learning. Despite successful induction and maintenance of synaptic plasticity, we found deficits of memory consolidation when there were defects in intrinsic plasticity. Our results suggest that intrinsic plasticity of cerebellar Purkinje cell has a significant role in motor memory consolidation.

Long-term depression (LTD) at the parallel fiber (PF)-to-cerebellar Purkinje cell (PC) synapse is implicated in the output of PCs, the sole output of the cerebellar cortex. In addition to synaptic plasticity, intrinsic excitability is also one of the components that determines PC output. Although long-term potentiation of intrinsic excitability (LTP-IE) has been suggested, it has yet to be investigated how PF–PC LTD modifies intrinsic excitability of PCs. Here, we show that pairing of the PF and climbing fiber (CF) for PF–PC LTD induction evokes LTD-IE in cerebellar PCs from male C57BL/6 mice. Interestingly, this intrinsic plasticity showed different kinetics from synaptic plasticity, but both forms of plasticity share Ca²⁺ signaling and protein kinase C pathway as their underlying mechanism. Although small-conductance Ca²⁺-activated K⁺ channels play important roles in LTP-IE, no direct implication has been found. After PF–PC LTD induction, neither the temporal summation of dendritic EPSP nor the power of spike frequency adaptation is changed, indicating that cerebellar LTD executes the information processing in a quantitative way without quality changes of synaptic integration and generation of output signals. Our results suggest that LTD-IE may have a synergistic effect with synaptic depression on the total net output of neurons by amplifying the modification of PF synaptic transmission. SIGNIFICANCE STATEMENT Although the output of Purkinje cells (PCs) is a critical component of cerebellum-dependent learning and memory, the changes of PC excitability when synaptic LTD occurs are unclear. Here, we show that the induction of PF–PC LTD evokes LTD-IE in PCs. Our observation complements previous intrinsic plasticity phenomenon of long-term potentiation of intrinsic excitability (LTP-IE), providing evidence for the idea that intrinsic plasticity has bidirectionality as synaptic plasticity. LTD-IE occurs together with synaptic LTD and both phenomena are dependent on the Ca²⁺ signaling pathway. Furthermore, our findings raise the prospect that this synaptic and intrinsic plasticity acts synergistically in PCs to modify neuronal activity in the same direction when learning occurs.

Vascular cognitive impairment (VCI) is a common sequela of cerebrovascular disorders. Although transcutaneous auricular vagus nerve stimulation (taVNS) has been considered a complementary treatment for various cognitive disorders, preclinical data on the effect of taVNS on VCI and its mechanism remain ambiguous. To measure cerebrospinal fluid (CSF) circulation during taVNS, we used in vivo two-photon microscopy with CSF and vasculature tracers. VCI was induced by transient bilateral common carotid artery occlusion (tBCCAO) surgery in mice. The animals underwent anesthesia, off-site stimulation, or taVNS for 20 min. Cognitive tests, including the novel object recognition and the Y-maze tests, were performed 24 h after the last treatment. The long-term treatment group received 6 days of treatment and was tested on day 7; the short-term treatment group received 2 days of treatment and was tested 3 days after tBCCAO surgery. CSF circulation increased remarkably in the taVNS group, but not in the anesthesia-control or off-site-stimulation-control groups. The cognitive impairment induced by tBCCAO was significantly restored after both long- and short-term taVNS. In terms of effects, both long- and short-term stimulations showed similar recovery effects. Our findings provide evidence that taVNS can facilitate CSF circulation and that repetitive taVNS can ameliorate VCI symptoms.

Group I metabotropic glutamate receptors (mGlu1/5 receptors) have important roles in synaptic activity in the central nervous system. They modulate neuronal excitability by mobilizing intracellular Ca2+ following receptor activation. Also, accumulating evidence has indicated the association of Ca2+ signaling with lipid rafts. Caveolin, an adaptor protein found in a specialized subset of lipid rafts, has been reported to promote the localization of membrane proteins to lipid rafts.

Intrinsic plasticity of cerebellar Purkinje cells (PCs) is recently highlighted in the cerebellar local circuits, however, its physiological impact on the cerebellar learning and memory remains elusive. Using a mouse model of memory consolidation deficiency, we found that the intrinsic plasticity of PCs may be involved in motor memory consolidation. Gain-up training of the vestibulo-ocular reflex produced a decrease in the synaptic weight of PCs in both the wild-type and knockout groups. However, intrinsic plasticity was impaired only in the knockout mice. Furthermore, the observed defects in the intrinsic plasticity of PCs led to the formation of improper neural plasticity in the vestibular nucleus (VN) neurons. Our results suggest that the synergistic modulation of intrinsic and synaptic plasticity in PCs is required for the changes in following plasticity in the VN, thereby contributes to the long-term storage of motor memory.