Here we investigate stimulus generalization in a cerebellar learning paradigm, called eyeblink conditioning. Mice were conditioned to close their eyes in response to a 10 kHz tone by repeatedly pairing this tone with an air puff to the eye 250 ms after tone onset. After ten consecutive days of training, when mice showed reliable conditioned eyelid responses to the 10 kHz tone, we started to expose them to tones with other frequencies, ranging from 2 to 20 kHz. We found that mice had a strong generalization gradient, whereby the probability and amplitude of conditioned eyelid responses gradually decreases depending on the dissimilarity with the 10 kHz tone. Tones with frequencies closest to 10kHz evoked the most and largest conditioned eyelid responses and each step away from the 10 kHz tone resulted in fewer and smaller conditioned responses. In addition, we found that tones with lower frequencies resulted in conditioned responses that peaked earlier after tone onset compared to those to tones with higher frequencies. Together, our data show prominent generalization patterns in cerebellar learning. Since the known function of cerebellum is rapidly expanding from pure motor control to domains that include cognition, reward-learning, fear-learning, social function, and even addiction, our data implies generalization controlled by cerebellum in all these domains. Significance Statement: Conditioned stimuli that resemble each other will evoke a rather similar behavioral response. Here, we investigate this phenomenon of stimulus generalization using Pavlovian eyeblink conditioning to probe cerebellar function. Cerebellum is a brain structure whose known function is rapidly expanding from pure motor control to domains that include cognition, reward-learning, fear-learning, social function, and even addiction. Since we found a strong generalization of the auditory stimulus in eyeblink conditioning, our data implies an important role for generalization in motor and non-motor domains. Methods SubjectsWe used 14 wild-type C57Bl/6 mice, 7 of which were male and 7 were female. Mice were between 11-16 weeks old at the start of the experiment. All mice were housed individually during the experiment with food and water ad libitum in a 12:12 light/dark cycle. Experiments were performed during the light phase. All experiments were performed in accordance with the European Communities Council Directive. Protocols were reviewed and approved by the Erasmus Laboratory Animal Science Center (work protocol nr. 15-273-138; project license nr. AVD101002015273) Auditory Brainstem Responses Since C57Bl/6 mice are prone for developing hearing problems, we recorded prior to the start of eyeblink conditioning training the auditory brainstem responses (ABRs) to obtain hearing level thresholds. Mice were anesthetized with a ketamine/xylazine mixture at a dose of 100/10 mg/kg body weight, administered intraperitoneally (ketamine: Alfasan, Woerden, NL; xylazine: Sedazine®, AST Farma, Oudewater, NL). After this, they were placed in a soundand ligh...

Individuals with Neurofibromatosis type 1 (NF1) experience a high degree of motor problems. The cerebellum plays a pivotal role in motor functioning and the NF1 gene is highly expressed in cerebellar Purkinje cells. However, it is not well understood to what extent NF1 affects cerebellar functioning and how this relates to NF1 motor functioning. Therefore, we subjected global Nf1+/− mice to a cerebellum-dependent associative learning task, called Pavlovian eyeblink conditioning. Additionally, we assessed general motor function and muscle strength in Nf1+/− mice. To our surprise, we found that Nf1+/− mice showed a moderately increased learning rate of conditioned eyeblink responses, as well as improved accuracy in the adaptive timing of the eyeblink responses. Locomotion, balance, general motor function, and muscle strength were not affected in Nf1+/− mice. Together, our results support the view that cerebellar function in Nf1+/− mice is unimpaired.

Tests of human brain circuit function typically require fixed equipment in lab environments. We have developed a smartphone-based platform for neurometric testing. This platform, which uses AI models like computer vision, is optimized for at-home use and produces reproducible, robust results on a battery of tests, including eyeblink conditioning, prepulse inhibition of acoustic startle response, and startle habituation. This approach provides a scalable, universal resource for quantitative assays of central nervous system function.