Instructive climbing fiber signals outside the cerebellum - The classic example of supervised learning occurs at the parallel fiber (PF) to Purkinje cell synapse in the cerebellum, where plasticity depends on co-activity of the climbing fiber (CF) input, which – according to the theories of Marr, Albus and Ito – provides the error signal. In a wider interpretation, this signal is instructive in nature, and might be related to error, sensory omission, as well as reward or reward-prediction. Depending on the proper timing intervals between the two stimuli, CF co-activity with the PF input promotes synaptic long- term depression (LTD) at PF synapses and thus helps to optimize synaptic input weights. This well-studied function of CFs within the cerebellar system is in stark contrast to what is known about the potential relevance of CF activity outside of the cerebellum. A plausible anatomical pathway for various interactions with neocortical areas has been described, which includes activation of Purkinje cells, and the subsequent signal transfer via cerebellar nuclei and thalamic nuclei, e.g. the ventral lateral (VL) and posteriormedial nuclei (Pom). Here, we ask whether CF co-activity can provide an instructive signal that affects receptive field (RF) plasticity in the primary somatosensory (S1; barrel) cortex of mice. Preliminary data from our laboratory show that repetitive activation of individual whiskers enhances whisker representation in the barrel cortex as assessed by intrinsic optical imaging. Optogenetic co-activation of channelrhodopsin 2 (ChR2)-expressing CFs in the cerebellum suppresses this form of cortical RF plasticity. These data show that indeed CF signaling may act as an instructive signal for plasticity outside of the cerebellum and supervises learning in the neocortex. However, intrinsic imaging does not provide information about participating cellular structures, e.g. which neurons change in this form of RF plasticity and which neurons ultimately mediate the effects of CF activity. In this study, we propose to use two-photon microscopy and optogenetics in awake mice to address the following questions. First, we will assess how whisker stimulation affects S1 cortex circuitry (aim 1). We will test the hypothesis that the RF plasticity observed with intrinsic imaging is due to an increase in the activity of L2/3 pyramidal neurons. We will also zoom in on parvalbumin-expressing (PV+) interneurons to follow up on our pilot data that show that these PV+ interneurons downregulate their activity after whisker tetanization. Second, we will examine how optogenetic co-activation of CF terminals in the cerebellum with 470nm light pulses impairs activity changes in these neuronal populations (aim 2). Next to pyramidal neurons, our focus will again be on PV+ interneurons, as our pilot data show that optogenetic CF stimulation drives activation of these inhibitory neurons, thus providing a potential pathway for the observed impact on the cortical network. Third, we will study whether direct optogenetic Purkinje cell activation mimics the effects of CF activation, addressing the question whether CFs act via the cerebellar cortex or a direct effect on the cerebellar nuclei (aim 3). This work will be the first to investigate whether CF signaling has an instructive role beyond the cerebellum.
