Expression of Protein Kinase C Inhibitor Blocks Cerebellar Long-Term Depression without Affecting Purkinje Cell Excitability in Alert Mice
Résumé
A longstanding but still controversial hypothesis is that long-term depression (LTD) of parallel fiber-Purkinje cell synapses in the cerebellum embodies part of the neuronal information storage required for associative motor learning. Transgenic mice in which LTD is blocked by Purkinje cell-specific inhibition of protein kinase C (PKC) (L7-PKCI mutants) do indeed show impaired adaptation of their vestibulo-ocular reflex, whereas the dynamics of their eye movement performance are unaffected. However, because L7-PKCI mutants have a persistent multiple climbing fiber innervation at least until 35 d of age and because the baseline discharge of the Purkinje cells in the L7-PKCI mutants is unknown, factors other than a blockage of LTD induction itself may underlie their impaired motor learning. We therefore investigated the spontaneous discharge of Pur-kinje cells in alert adult L7-PKCI mice as well as their multiple climbing fiber innervation beyond the age of 3 months. We found that the simple spike and complex spike-firing properties (such as mean firing rate, interspike interval, and spike count variability), oscillations, and climbing fiber pause in the L7-PKCI mutants were indistinguishable from those in their wild-type littermates. In addition, we found that multiple climbing fiber innervation does not occur in cerebellar slices obtained from 3-to 6-month-old mutants. These data indicate (1) that neither PKC inhibition nor the subsequent blockage of LTD induction disturbs the spontaneous discharge of Purkinje cells in alert mice, (2) that Purkinje cell-specific inhibition of PKC detains rather than prevents the developmental conversion from multiple to mono-innervation of Purkinje cells by climbing fibers, and (3) that as a consequence the impaired motor learning as observed in older adult L7-PKCI mutants cannot be attributable either to a disturbance in the baseline simple spike and complex spike activities of their Purkinje cells or to a persistent multiple climbing fiber innervation. We conclude that cerebellar LTD is probably one of the major mechanisms underlying motor learning, but that deficits in LTD induction and motor learning as observed in the L7-PKCI mutants may only be reflected in differences of the Purkinje cell signals during and/or directly after training. A challenge faced by neuroscience is to understand how model systems of information storage in the brain, such as long-term potentiation and long-term depression (LTD), function in neural circuits that control behavioral learning. Cerebellar LTD is an attenuation of the granule cell axon-Purkinje cell synapse that occurs after conjunctive stimulation of the granule cell axons and climbing fiber inputs (Ito et al., 1982; Linden and Connor, 1995). It has been suggested that LTD underlies several forms of motor learning, including adaptation of the vestibulo-ocular reflex (VOR) and eye blink conditioning (Ito, 1989). Several studies using knockout mice have supported this claim by showing correlations between deficits in LTD and behavioral learning (Aiba et al., 1994; Conquet et al., 1994; Funabiki et al., 1995; Kashiwabuchi et al., 1995; Shibuki et al., 1996). However, the lack of anatomical specificity and functional compensation via similar gene family members has complicated the analyses of knockout mice. For example, global knockouts of the type I metabotropic glutamate receptor (mGluR1) show a blockade of LTD induction and impaired eye blink conditioning (Aiba et al., 1994; Conquet et al., 1994), but because this receptor is expressed at multiple sites in the brain, it is difficult to correlate the physiological and behavioral phenotype of the cell (cf. Ichise et al., 2000). To overcome these limitations, we have created a trans-genic mouse in which a protein kinase C (PKC) inhibitory pep-tide, PKC[19-31], is selectively expressed in Purkinje cells (De Zeeuw et al., 1998). Cultured Purkinje cells from these L7-PKCI mice show a complete blockade of LTD induction, and behavioral analysis indicates a loss of VOR adaptation, whereas the default
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