Purkinje Cell Collaterals Preferentially Target a Subtype of Molecular Layer Interneuron

Area of Science:

• Neuroscience
• Cellular Neuroscience
• Cerebellar Circuitry

Background:

• Purkinje cells (PCs) are key output neurons in the cerebellar cortex, projecting to other PCs, molecular layer interneurons (MLIs), and Purkinje layer interneurons (PLIs).
• The function of PC collateral synapses onto MLIs was previously assumed to provide positive feedback to PCs, based on the premise that MLIs primarily inhibit PCs.
• Recent discoveries identified two MLI subtypes: MLI1s inhibit PCs, while MLI2s inhibit MLI1s and disinhibit PCs, necessitating a re-evaluation of PC-MLI pathway influence.

Purpose of the Study:

• To elucidate the precise connectivity of Purkinje cell (PC) axon collaterals onto the identified subtypes of molecular layer interneurons (MLIs).
• To understand the functional implications of PC-MLI subtype connectivity for feedback regulation within the cerebellar cortex.
• To clarify the role of PC-MLI synapses in the context of other PC feedback pathways, including PC-PC and PC-PLI connections.

Main Methods:

• Serial electron microscopy (EM) was employed to reconstruct and quantify synaptic connections between PCs and MLI subtypes in mice.
• Optogenetic stimulation of PCs was performed in slice experiments to record evoked currents in MLI subtypes, assessing functional synaptic effects.
• Synaptic input from candelabrum cells (a type of PLI) onto MLI subtypes was also characterized.

Main Results:

• EM reconstructions revealed that PC axon collaterals form synapses onto PCs (53%), PLIs (32%), MLI1s (6%), and MLI2s (7%).
• Despite lower synapse numbers, MLI2s receive significantly more PC synapses per neuron than MLI1s due to their lower abundance.
• Optogenetic activation of PCs evoked inhibitory currents in MLI2s and disinhibitory effects in MLI1s, confirming functional differentiation. Candelabrum cells preferentially synapse onto MLI1s.

Conclusions:

• PC-MLI synapses do not primarily mediate disinhibition of PCs; instead, they preferentially inhibit MLI2s and disinhibit MLI1s.
• The PC-MLI2-MLI1-PC and PC-PLI-MLI1-PC pathways, along with direct PC-PC synapses, likely contribute to negative feedback, counteracting increases in PC firing.
• These findings refine our understanding of cerebellar circuit dynamics and the complex regulatory roles of Purkinje cell feedback.