Calcium dynamics following INS were also investigated in the mouse cerebellum. Previous studies have used flavin autofluorescence imaging, an indirect measure of the rate of mitochondrial ATP production based on the redox state of flavin molecules, such as flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN),17 to assess neuronal activity in the cerebellum.21 Due to the neuronal architecture of the cerebellum, which is characterized by long parallel fibers lining the cortical folia, superficial electrical stimulation produces a long-narrow strip of flavin fluorescence and induces field potentials within the strip (beam) of parallel fibers stimulated. Lateral spread (perpendicular to parallel fibers) of the signal is prevented by action of inhibitory interneurons. Previous studies have shown that visualization of the activation along the parallel fibers by flavin imaging requires long electrical stimulation pulse trains (several seconds), presumably related to the time course of mitochondrial activation. However, similarly long pulses of INS do not lead to beam-like activation, but rather to diffuse activation at the stimulation site and is not correlated with the direction of parallel fibers. This diffuse response is eliminated with application of fluoroacetate, an astrocytic poison, suggesting that the activation induced by long train INS is astrocytic in nature. In contrast, stimulation with short INS pulse trains, even just a single optical pulse, results in neural activation, as evidenced by field potential recordings similar to the ones obtained using electrical stimulation. Consistent with this, calcium imaging following INS reveals a fairly focal region of activation, which is detectable at much lower pulse train durations () and is insensitive to fluoroacetate. This suggests that INS of short duration, a duration that is too brief to produce detectable changes in flavin fluorescence, activates neuronal response, which is detectable both electrophysiologically and with calcium imaging. Longer INS stimulation leads to accumulation of heat in the tissue, which leads to a delayed but significant astrocytic response. Therefore, the fact that INS can reveal the neural architecture inherent in cerebellar cortex confirms its spatial precision and usefulness as a tool for studying cortical circuitry. Moreover, the markedly different effects of short and long INS pulse trains underscore the importance of choice of stimulation parameters.