Cerebellar cortex involvement in classical eyelid conditioning

A model for cerebellar involvement in motor learning was tested using classical eyelid conditioning in the rabbit. Briefly, we assume that modifications of the strength of granule cell synapses at Purkinje cells in the cerebellar cortex and mossy fiber (MF) synapses at cerebellar interpositus nuclei are responsible for the acquisition, adaptively-timed expression, and extinction of conditioned eyelid responses (CRs). A corollary of these assumptions is that the cerebellar cortex is necessary for acquisition and extinction. This model also suggests a mechanism whereby the cerebellar cortex can discriminate different times during a conditioned stimulus (CS) and thus mediate the learned timing of CRs. Therefore, experiments were done to determine the role of the cerebellar cortex in the timing, extinction, and acquisition of CRs. Lesions of the cerebellar cortex that included the anterior lobe disrupted the learned timing of CRs such that they occurred at extremely short latencies. Stimulation of MFs in the middle cerebellar peduncle as the CS could support differently timed CRs in the same animal. These data indicate that synaptic plasticity in the cerebellar cortex mediates the learned timing of CRs. These short-latency CRs which resulted from anterior lobe damage did not extinguish, while CRs in animals receiving lesions which did not include the anterior lobe extinguished normally. Preliminary data suggests that lesions of the anterior lobe which produce short-latency responses prevent the acquisition of CRs to a novel CS. These findings indicate that the anterior lobe of cerebellar cortex is necessary for eyelid conditioning. The involvement of the anterior lobe in eyelid conditioning has not been previously reported, however, the anterior lobe has generally been spared in lesion studies examining cerebellar cortex involvement in eyelid conditioning due to its relatively inaccessible location. The observation that the anterior lobe of the cerebellar cortex is not always required for the basic expression of CRs, but is necessary for response timing, extinction, and acquisition, is consistent with the hypothesis that eyelid conditioning can involve plasticity in both the cerebellar cortex and interpositus nucleus and that plasticity in the nucleus is controlled by Purkinje cell activity. ^

Cerebellar mechanisms for motor learning: Testing predictions from a large-scale computer simulation

The cerebellum is the major brain structure that contributes to our ability to improve movements through learning and experience. We have combined computer simulations with behavioral and lesion studies to investigate how modification of synaptic strength at two different sites within the cerebellum contributes to a simple form of motor learning—Pavlovian conditioning of the eyelid response. These studies are based on the wealth of knowledge about the intrinsic circuitry and physiology of the cerebellum and the straightforward manner in which this circuitry is engaged during eyelid conditioning. Thus, our simulations are constrained by the well-characterized synaptic organization of the cerebellum and further, the activity of cerebellar inputs during simulated eyelid conditioning is based on existing recording data. These simulations have allowed us to make two important predictions regarding the mechanisms underlying cerebellar function, which we have tested and confirmed with behavioral studies. The first prediction describes the mechanisms by which one of the sites of synaptic modification, the granule to Purkinje cell synapses (gr → Pkj) of the cerebellar cortex, could generate two time-dependent properties of eyelid conditioning—response timing and the ISI function. An empirical test of this prediction using small, electrolytic lesions of the cerebellar cortex revealed the pattern of results predicted by the simulations. The second prediction made by the simulations is that modification of synaptic strength at the other site of plasticity, the mossy fiber to deep nuclei synapses (mf → nuc), is under the control of Purkinje cell activity. The analysis predicts that this property should confer mf → nuc synapses with resistance to extinction. Thus, while extinction processes erase plasticity at the first site, residual plasticity at mf → nuc synapses remains. The residual plasticity at the mf → nuc site confers the cerebellum with the capability for rapid relearning long after the learned behavior has been extinguished. We confirmed this prediction using a lesion technique that reversibly disconnected the cerebellar cortex at various stages during extinction and reacquisition of eyelid responses. The results of these studies represent significant progress toward a complete understanding of how the cerebellum contributes to motor learning. ^

CHARACTERIZATION AND TREATMENT OF A NOVEL MOUSE MODEL OF TSC-ASSOCIATED AUTISM

Tuberous sclerosis complex (TSC) is a dominant tumor suppressor disorder caused by mutations in either TSC1 or TSC2. The proteins of these genes form a complex to inhibit the mammalian target of rapamycin complex 1 (mTORC1), which controls protein translation and cell growth. TSC causes substantial neuropathology, often leading to autism spectrum disorders (ASDs) in up to 60% of patients. The anatomic and neurophysiologic links between these two disorders are not well understood. However, both disorders share cerebellar abnormalities. Therefore, we have characterized a novel mouse model in which the Tsc2 gene was selectively deleted from cerebellar Purkinje cells (Tsc2f/-;Cre). These mice exhibit progressive Purkinje cell degeneration. Since loss of Purkinje cells is a well-reported postmortem finding in patients with ASD, we conducted a series of behavior tests to assess if Tsc2f/-;Cre mice displayed autistic-like deficits. Using the three chambered social choice assay, we found that Tsc2f/-;Cre mice showed behavioral deficits, exhibiting no preference between a stranger mouse and an inanimate object, or between a novel and a familiar mouse. Tsc2f/-;Cre mice also demonstrated increased repetitive behavior as assessed with marble burying activity. Altogether, these results demonstrate that loss of Tsc2 in Purkinje cells in a haploinsufficient background lead to behavioral deficits that are characteristic of human autism. Therefore, Purkinje cells loss and/or dysfunction may be an important link between TSC and ASD. Additionally, we have examined some of the cellular mechanisms resulting from mutations in Tsc2 leading to Purkinje cell death. Loss of Tsc2 led to upregulation of mTORC1 and increased cell size. As a consequence of increased protein synthesis, several cellular stress pathways were upregulated. Principally, these included altered calcium signaling, oxidative stress, and ER stress. Likely as a consequence of ER stress, there was also upregulation of ubiquitin and autophagy. Excitingly, treatment with an mTORC1 inhibitor, rapamycin attenuated mTORC1 activity and prevented Purkinje cell death by reducing of calcium signaling, the ER stress response, and ubiquitin. Remarkably, rapamycin treatment also reversed the social behavior deficits, thus providing a promising potential therapy for TSC-associated ASD.