How Purkinje cell (PC) activity may be altered by learning is central to theories of the cerebellum. Pavlovian eyelid conditioning, because of how directly it engages the cerebellum, has helped reveal many aspects of cerebellar learning and the underlying mechanisms. Theories of cerebellar learning assert that climbing fiber inputs control plasticity at synapses onto PCs, and thus PCs control the expression of learned responses. We tested this assertion by recording 184 eyelid PCs and 240 non-eyelid PCs during the expression of conditioned eyelid responses (CRs) in well trained rabbits. By contrasting the responses of eyelid and non-eyelid PCs and by contrasting the responses of eyelid PCs under conditions that produce differently timed CRs, we test the hypothesis that learning-related changes in eyelid PCs contribute to the learning and adaptive timing of the CRs. We used a variety of analyses to test the quantitative relationships between eyelid PC responses and the kinematic properties of the eyelid CRs. We find that the timing of eyelid PC responses varies systematically with the timing of the behavioral CRs and that there are differences in the magnitude of eyelid PC responses between larger-CR, smaller-CR, and non-CR trials. However, eyelid PC activity does not encode any single kinematic property of the behavioral CRs at a fixed time lag, nor does it linearly encode CR amplitude. Even so, the results are consistent with the hypothesis that learning-dependent changes in PC activity contribute to the adaptively timed expression of conditioned eyelid responses.

The neural pathways that convey conditioned stimulus (CS) information to the cerebellum during eyeblink conditioning have not been fully delineated. It is well established that pontine mossy fiber inputs to the cerebellum convey CS-related stimulation for different sensory modalities (e.g., auditory, visual, tactile). Less is known about the sources of sensory input to the pons that are important for eyeblink conditioning. The first experiment of the current study was designed to determine whether electrical stimulation of the medial auditory thalamic nuclei is a sufficient CS for establishing eyeblink conditioning in rats. The second experiment used anterograde and retrograde tract tracing techniques to assess neuroanatomical connections between the medial auditory thalamus and pontine nuclei. Stimulation of the medial auditory thalamus was a very effective CS for eyeblink conditioning in rats, and the medial auditory thalamus has direct ipsilateral projections to the pontine nuclei. The results suggest that the medial auditory thalamic nuclei and their projections to the pontine nuclei are components of the auditory CS pathway in eyeblink conditioning.A primary emphasis in neurobiological analyses of Pavlovian eyeblink conditioning has been on identifying the anatomical sites and cellular mechanisms of memory storage (Christian and Thompson 2003;Thompson 2005). Less emphasis has been placed on identifying the neural pathways that conduct stimulation from conditioned stimuli to the sites of memory induction and retention. The cerebellum is the anatomical site of memory storage in Pavlovian eyeblink (eyelid and nictitating membrane movement) conditioning (Christian and Thompson 2003;Thompson 2005;Ohyama et al. 2006). Cerebellar function may be influenced by processes occurring within its sensory input pathways, interactions among components of the input pathways, and sources of feedback to the input pathways (Clark et al. 1997;Bao et al. 2000;Medina et al. 2002). A full characterization of the sensory input pathways to the cerebellum necessary for motor learning is, therefore, critical for developing a comprehensive understanding of cerebellar function. Identification of sensory input pathways to the cerebellum is also critical for elucidating the mechanisms underlying the ontogeny of motor learning (Freeman et al. 2005).Several key components of the conditioned stimulus (CS) pathway in eyeblink conditioning have been identified using lesion, inactivation, unit recording, stimulation, and tract tracing techniques (Steinmetz et al. 1986(Steinmetz et al. , 1987(Steinmetz et al. , 1989Lewis et al. 1987;Knowlton and Thompson 1988;Steinmetz 1990;Steinmetz and Sengelaub 1992;Gould et al. 1993;Tracy et al. 1998;Hesslow et al. 1999;Bao et al. 2000;Freeman and Rabinak 2004;Freeman et al. 2005). The pontine mossy fiber projection to the cerebellum is necessary for eyeblink conditioning with CSs of different modalities and stimulation of the pontine nuclei or the middle cerebellar peduncle is sufficient for eyeblink condit...

The neural mechanisms underlying excitatory and inhibitory eyeblink conditioning were compared using muscimol inactivation of the cerebellum. In experiment 1, rats were given saline or muscimol infusions into the anterior interpositus nucleus ipsilateral to the conditioned eye before each of four daily excitatory conditioning sessions. Postinfusion testing continued for four more excitatory conditioning sessions. All rats were given a final test session after muscimol infusions. The muscimol infusions inactivated the cerebellar nuclei, lateral anterior lobe, crus I, rostral crus II, and lobule HVI ipsilateral to the conditioned eye. Acquisition of excitatory conditioning was completely prevented by muscimol inactivation. In experiment 2, there were four experimental phases. Phase 1 consisted of excitatory conditioning. In phase 2, rats were given saline or muscimol infusions before conditioned inhibition training. Phase 3 consisted of continued conditioned inhibition training with no drug infusions. In phase 4, all rats received a retardation test in which the inhibitory stimulus was paired with the unconditioned stimulus. Muscimol infusions blocked the expression of conditioned responses during phase 2. However, robust conditioned inhibition was evident in phases 3 and 4. The findings indicate that conditioned excitation and inhibition depend on different mechanisms.

The auditory conditioned stimulus (CS) pathway that is necessary for delay eyeblink conditioning was investigated using reversible inactivation of the medial auditory thalamic nuclei (MATN) consisting of the medial division of the medial geniculate (MGm), suprageniculate (SG), and posterior intralaminar nucleus (PIN). Rats were given saline or muscimol infusions into the MATN contralateral to the trained eye before each of four conditioning sessions with an auditory CS. Rats were then given four additional sessions without infusions to assess savings from the initial training. All rats were then given a retention test with a muscimol infusion followed by a recovery session. Muscimol infusions through cannula placements within 0.5 mm of the MGm prevented acquisition of eyeblink conditioned responses (CRs) and also blocked CR retention. Cannula placements more than 0.5 mm from the MATN did not completely block CR acquisition and had a partial effect on CR retention. The primary and secondary effects of MATN inactivation were examined with 2-deoxy-glucose (2-DG) autoradiography. Differences in 2-DG uptake in the auditory thalamus were consistent with the cannula placements and behavioral results. Differences in 2-DG uptake were found between groups in the ipsilateral auditory cortex, basilar pontine nuclei, and inferior colliculus. Results from this experiment indicate that the MATN contralateral to the trained eye and its projection to the pontine nuclei are necessary for acquisition and retention of eyeblink CRs to an auditory CS.Pavlovian eyeblink conditioning is an ideal paradigm for identifying the necessary and sufficient neural structures underlying associative learning (Thompson 1976(Thompson , 1983. Eyeblink conditioning is typically established by pairing a conditioned stimulus (CS) such as a tone or light with an unconditioned stimulus (US) that elicits the eyeblink reflex. An eyeblink conditioned response (CR) to the CS emerges over the course of training and occurs prior to the onset of the US. Permanent lesions, reversible inactivation, stimulation, unit recording, and neuroanatomy have been used to establish that the interpositus nucleus and cerebellar cortex are the necessary sites of learning-dependent plasticity underlying eyeblink conditioning (Thompson 2005). The basilar pontine nuclei are the proximal end of the CS pathway, which send mossy fiber input to the contralateral cerebellar cortex and interpositus nucleus (Steinmetz et al. 1986(Steinmetz et al. , 1987Lewis et al. 1987;Steinmetz 1990;Steinmetz and Sengelaub 1992;Hesslow et al. 1999;Freeman and Rabinak 2004;Freeman et al. 2005).Possible auditory inputs to the pontine nuclei that could provide auditory CS information have been proposed (Steinmetz et al. 1987;Steinmetz and Sengelaub 1992), but a systematic experimental analysis of these inputs has not been done. The ventral cochlear nucleus has a monosynaptic projection to the pontine nuclei in rats and rabbits (Steinmetz et al. 1987;Campolattaro et al. 2007), but this short-latency audit...

Eyeblink conditioning, a type of associative motor learning, requires the cerebellum. The medial auditory thalamus is a necessary source of stimulus input to the cerebellum during auditory eyeblink conditioning. Nothing is currently known about interactions between the thalamus and cerebellum during associative learning. In the current study, neuronal activity was recorded in the cerebellar interpositus nucleus and medial auditory thalamus simultaneously from multiple tetrodes during auditory eyeblink conditioning to examine the relative timing of learning-related plasticity within these interconnected areas. Learning-related changes in neuronal activity correlated with the eyeblink conditioned response were evident in the cerebellum before the medial auditory thalamus over the course of training and within conditioning trials, suggesting that thalamic plasticity may be driven by cerebellar feedback. Short-latency plasticity developed in the thalamus during the first conditioning session and may reflect attention to the conditioned stimulus. Extinction training resulted in a decrease in learning-related activity in both structures and an increase in inhibition within the cerebellum. A feedback projection from the cerebellar nuclei to the medial auditory thalamus was identified, which may play a role in learning by facilitating stimulus input to the cerebellum via the thalamo-pontine projection.