Marlies Oostland

High-density probes allow electrophysiological recordings from many neurons simultaneously across entire brain circuits but don’t reveal cell type. Here, we develop a strategy to identify cell types from extracellular recordings in awake animals, revealing the computational roles of neurons with distinct functional, molecular, and anatomical properties. We combine optogenetic activation and pharmacology using the cerebellum as a testbed to generate a curated ground-truth library of electrophysiological properties for Purkinje cells, molecular layer interneurons, Golgi cells, and mossy fibers. We train a semi-supervised deep-learning classifier that predicts cell types with greater than 95% accuracy based on waveform, discharge statistics, and layer of the recorded neuron. The classifier’s predictions agree with expert classification on recordings using different probes, in different laboratories, from functionally distinct cerebellar regions, and across animal species. Our classifier extends the power of modern dynamical systems analyses by revealing the unique contributions of simultaneously-recorded cell types during behavior.

Summary: Perturbation to the cerebellum can lead to deficits in motor function, cognition, and behavioral flexibility. Here we report that a cerebellum-specific transgenic mouse autism model with disrupted Purkinje cell function shows unexpectedly accelerated learning on a sensory evidence-accumulation task, as well as enhanced sensory reactivity to touch and auditory cues. Computational latent-state analysis of behavior revealed that accelerated learning was associated with enhanced focus on current over past trials. In on-task states, a subset of Purkinje cells in crus I produced more complex spikes to sensory stimuli. Learning was accelerated by providing cue-locked optogenetic stimulation of Purkinje cells, but unaffected by continuous optogenetic interference with Purkinje cell activity. Complex spikes fired in response to both correct and incorrect choices, but less so when mice were on-task. Both transgenic mice and mice receiving cue-locked optogenetic stimulation showed prolonged sensory responses in Purkinje-cell complex spikes and anterior cingulate cortex. We suggest that cerebellar activity may shape evidence-accumulation learning by enhancing task focus and neocortical processing of current experience. Highlights: Faster learning and enhanced sensory salience with cerebellar manipulations in mice Accelerated learning arises from prolonged occupancy in an on-task behavioral state Cerebellar manipulations can influence neocortex via altered complex spike activity Cerebellum findings consistent with a weak global coherence account of autism eTOC blurb: In a cerebellum-based mouse autism model, perturbed function leads to faster learning of a working-memory task, mediated by higher focus on current trials. The effects are emulated by optogenetic perturbation of Purkinje cells, and both perturbations drive enhanced neocortical activity. Results are consistent with a weak coherence model for autism.

To make successful evidence-based decisions, the brain must rapidly and accurately transform sensory inputs into specific goal-directed behaviors. Most experimental work on this subject has focused on forebrain mechanisms. Using a novel evidence-accumulation task for mice, we performed recording and perturbation studies of crus I of the lateral posterior cerebellum, which communicates bidirectionally with numerous forebrain regions. Cerebellar inactivation led to a reduction in the fraction of correct trials. Using two-photon fluorescence imaging of calcium, we found that Purkinje cell somatic activity contained choice/evidence-related information. Decision errors were represented by dendritic calcium spikes, which in other contexts are known to drive cerebellar plasticity. We propose that cerebellar circuitry may contribute to computations that support accurate performance in this perceptual decision-making task.

Serotonin plays an important role of controlling the physiology of the cerebellum. However, serotonin receptor expression has not been fully studied in the developing cerebellum. We have recently shown that cerebellar granule cells transiently express 5-HT₃ receptors. In the present study, we investigate expression of 5-HT₁ and 5-HT₂ receptors in the mouse cerebellum both during postnatal development and in juvenile mice. Here, we show for the first time that 5-HT_1A and 5-HT_2A receptors are present on cerebellar granule cells with a distinct temporal expression pattern: 5-HT_1A receptors are expressed only during the first 2 weeks, while 5-HT_2A receptor expression persists until at least 8 weeks after birth. Because of its prolonged expression pattern, we investigated the electrophysiological properties of the 5-HT_2A receptor. 5-HT_2A receptors expressed by cerebellar granule cells promote stability by reducing variability of the synaptic response, and they modulate the paired-pulse ratio of the parallel fibre–Purkinje cell synapse. Furthermore, pharmacological block of 5-HT_2A receptors enhances short-term synaptic plasticity at the parallel fibre–Purkinje cell synapse. We thus show a novel role for serotonin in controlling function of the cerebellum via 5-HT_2A receptors expressed by cerebellar granule cells.

Key points: Serotonin 3 (5‐HT₃) receptors are expressed by excitatory granule cells in the cerebellum during early postnatal development. Here we show a novel role for serotonin in the regulation of cerebellar postnatal development via 5‐HT₃ receptors. Using 5‐HT_3A receptor knockout mice we show that 5‐HT₃ receptors expressed by granule cells, via the glycoprotein reelin, regulate the morphological maturation of Purkinje cells. The 5‐HT_3A receptor knockout mice show abnormal physiological maturation of Purkinje cells and impaired short‐term plasticity at the parallel fibre–Purkinje cell synapse, resulting in delayed climbing fibre elimination. With these results, we provide a better understanding of the role of serotonin in the developing brain, the control it has on the postnatal maturation of the cerebellum, and the cerebellum as a highly adaptive system during early postnatal development. Abstract  Functional serotonin 3 (5‐HT₃) receptors are transiently expressed by cerebellar granule cells during early postnatal development, where they modulate short‐term synaptic plasticity at the parallel fibre–Purkinje cell synapse. Here, we show that serotonin controls maturation of Purkinje cells in the mouse cerebellum. The 5‐HT₃ receptors regulate morphological maturation of Purkinje cells during early postnatal development, and this effect is mediated by the glycoprotein reelin. Using whole‐cell patch‐clamp recordings we also investigated physiological development of Purkinje cells in 5‐HT_3A receptor knockout mice during early postnatal development, and found abnormal physiological maturation, characterized by a more depolarized resting membrane potential, an increased input resistance and the ability to fire action potentials upon injection of a depolarizing current at an earlier age. Furthermore, short‐term synaptic plasticity was impaired at both the parallel fibre–Purkinje cell and the climbing fibre–Purkinje cell synapses, and both the amplitude and the frequency of spontaneous miniature events recorded from Purkinje cells were increased. The expedited morphological and physiological maturation affects the whole cerebellar cortical network, as indicated by delayed climbing fibre elimination in 5‐HT_3A receptor knockout mice. There was no difference between wild‐type and 5‐HT_3A receptor knockout mice in any of the morphological or physiological properties described above at later ages, indicating a specific time window during which serotonin regulates postnatal development of the cerebellum via 5‐HT₃ receptors expressed by granule cells.

The serotonin 5-HT(3) receptor is the only ligand-gated ion channel activated by serotonin and is expressed by GABAergic interneurons in many brain regions, including the cortex, amygdala and hippocampus. Furthermore, 5-HT(3) receptors are expressed by glutamatergic Cajal-Retzius cells in the cerebral cortex. We used 5-HT(3A)/enhanced green fluorescent protein (EGFP) transgenic mice to show that 5-HT(3) receptors are also ubiquitously expressed by glutamatergic granule cells in the cerebellum during the first three postnatal weeks. Using whole-cell patch clamp recordings, we show that local application of either serotonin or the selective 5-HT(3) receptor agonist SR57227A to granule cells results in a small inward current, demonstrating a post- and/or extrasynaptic localisation of the 5-HT(3) receptors. Functional 5-HT(3) receptors were also observed presynaptically at the parallel fibre-Purkinje cell synapse. Pharmacological block using the selective 5-HT(3) receptor antagonist tropisetron induced a reduction in the frequency of miniature synaptic events recorded from Purkinje cells. Paired-pulse stimulation of parallel fibres on whole-cell voltage clamped Purkinje cells from 1-week-old mice did not yet show synaptic plasticity. In the presence of tropisetron, the parallel fibre-Purkinje cell synapse showed paired-pulse depression. Taken together, these results show that functional 5-HT(3) receptors are present during early postnatal development in the cerebellum, where they modulate synaptic plasticity.