Diversity of cellular physiology and morphology of Purkinje cells in the adult zebrafish cerebellum

Abstract

This study was designed to explore the functional circuitry of the adult zebrafish cerebellum, focusing on its Purkinje cells and using whole-cell patch recordings and single cell labeling in slice preparations. Following physiological characterizations, the recorded single cells were labeled for morphological identification. It was found that the zebrafish Purkinje cells are surprisingly diverse. Based on their physiology and morphology, they can be classified into at least three subtypes: Type I, a narrow spike cell, which fires only narrow Na + spikes (<3 ms in duration), and has a single primary dendrite with an arbor restricted to the distal molecular layer; Type II, a broad spike cell, which fires broad Ca 2+ spikes (5-7 ms in duration) and has a primary dendrite with limited branching in the inner molecular layer and then further radiates throughout the molecular layer; and Type III, a very broad spike cell, which fires very broad Ca 2+ spikes (≥10 ms in duration) and has a dense proximal dendritic arbor that is either restricted to the inner molecular layer (Type IIIa), or radiates throughout the entire molecular layer (Type IIIb). The graded paired-pulse facilitation of these Purkinje cells' responses to parallel fiber activations and the all-or-none, paired-pulse depression of climbing fiber activation are largely similar to those reported for mammals. The labeled axon terminals of these Purkinje cells end locally, as reported for larval zebrafish.The present study provides evidence that the corresponding functional circuitry and information processing differ from what has been well-established in the mammalian cerebellum.

Highly synchronized inhibition from Purkinje cells entrains cerebellar output in zebrafish

“…At least two molecularly distinct subpopulations of ECs have been identified: olig2 + and calb2b + ECs, each occupying distinct regions in the dorsoventral and mediolateral axes of the cerebellum, with the 156A gal4 line that we used in this study, labeling both olig2 + and olig2 - populations (McFarland et al, 2008; Bae et al, 2009; Takeuchi et al, 2015). Zebrafish ECs have also been shown to have diverse dendritic morphology and projection patterns and the projection targets of ECs correlate with their location within the cerebellum, suggesting a molecular code for the topographic organization of different EC types (Heap et al, 2013; Magnus et al, 2023). We do not yet know if the chatty and stuttering types we observe correspond to these molecularly and topographically distinct populations of ECs but such heterogeneity in EC populations will not be surprising.…”

“…DCNs are known to be heterogenous with respect to their projection patterns, molecular expression patterns, firing properties and transmitter phenotypes. While DCNs consist of glutamatergic, GABAergic and glycinergic neurons (Bagnall et al, 2009; Uusisaari and Knöpfel, 2012; Kebschull et al, 2024), no inhibitory subtypes have been identified among ECs in teleosts and they seem to be a fully glutamatergic, long range projecting neuronal class (Bae et al, 2009; Heap et al, 2013; Matsui et al, 2014; Takeuchi et al, 2015; Harmon et al, 2020; Magnus et al, 2023). Given the above, one might speculate that distinct EC populations in fish might be related to ancestral glutamatergic populations observed in amniotes.…”

“…DCN/ECs are spontaneously firing cells and receive high frequency excitatory and inhibitory inputs. While mammalian DCNs receive mossy fiber and climbing fiber excitatory inputs (McCrea et al, 1977; Shinoda and Sugihara, 2013), fish ECs receive granule cell excitatory inputs (Ikenaga et al, 2006, 2023; Magnus et al, 2023). Also, while mammalian DCNs receive local GABAergic inhibition from other DCNs, in addition to PN inhibition, fish ECs appear to receive only PN inhibition (Bae et al, 2009; Heap et al, 2013; Kebschull et al, 2020, 2024).…”

“…This implies that synchronization of simple spikes among PNs could trigger strong inhibition of ECs. Lastly, both mammalian DCNs and ECs of adult zebrafish exhibit rebound spiking (Jahnsen, 1986; Aizenman and Linden, 1999; Magnus et al, 2023), suggesting that strong, synchronized inhibition could trigger timed spikes in these cells.…”

“…In mammals, DCN receives direct inputs from mossy fibers and olivary climbing fibers (McCrea et al, 1977;Shinoda and Sugihara, 2013). In teleosts, ECs receive inputs from PFs of granule cells which are post-synaptic to mossy fibers and seem to not receive climbing fiber input (Ikenaga et al, 2006(Ikenaga et al, , 2023Magnus et al, 2023). While in mammals, the DCN neurons are located ventrally within the cerebellar white matter, in teleosts such as zebrafish, the ECs are located proximal to the Purkinje cell layer, making them easily accessible for in vivo recordings and manipulation.…”