Cerebellar Zonal Patterning Relies on Purkinje Cell Neurotransmission

Abstract

Cerebellar circuits are patterned into an array of topographic parasagittal domains called zones. The proper connectivity of zones is critical for motor coordination and motor learning, and in several neurological diseases cerebellar circuits degenerate in zonal patterns. Despite recent advances in understanding zone function, we still have a limited understanding of how zones are formed. Here, we focused our attention on Purkinje cells to gain a better understanding of their specific role in establishing zonal circuits. We used conditional mouse genetics to test the hypothesis that Purkinje cell neurotransmission is essential for refining prefunctional developmental zones into sharp functional zones. Our results show that inhibitory synaptic transmission in Purkinje cells is necessary for the precise patterning of Purkinje cell zones and the topographic targeting of mossy fiber afferents. As expected, blocking Purkinje cell neurotransmission caused ataxia. Using in vivo electrophysiology, we demonstrate that loss of Purkinje cell communication altered the firing rate and pattern of their target cerebellar nuclear neurons. Analysis of Purkinje cell complex spike firing revealed that feedback in the cerebellar nuclei to inferior olive to Purkinje cell loop is obstructed. Loss of Purkinje neurotransmission also caused ectopic zonal expression of tyrosine hydroxylase, which is only expressed in adult Purkinje cells when calcium is dysregulated and if excitability is altered. Our results suggest that Purkinje cell inhibitory neurotransmission establishes the functional circuitry of the cerebellum by patterning the molecular zones, fine-tuning afferent circuitry, and shaping neuronal activity.

In vivo analysis of Purkinje cell firing properties during postnatal mouse development

“…In the future, it would be interesting to use juxtacellular labeling (Pinault 1996) to anatomically identify recorded neurons that were developmentally "marked" based on their genetic lineage (Sillitoe et al 2009) and then examine whether each class of Purkinje cell, based on its firing pattern, has a distinct location in the cerebellum, specific afferent connectivity, and unique developmental fate within the zonal "stripe" expression map (Sillitoe and Joyner 2007;White and Sillitoe 2013a). Indeed, the formation of zones is critically dependent on proper Purkinje cell firing (White et al 2014).…”

“…Unfortunately, we have a limited understanding of how motor behavior arises during development (Jacquelin et al 2012). In the future, it would be interesting to test whether the changes in firing during development indeed influence the relationship between spike types and if an increased interdependence of simple spikes and complex spikes correlates with improved motor control (Cerminara and Rawson 2004;Chaumont et al 2013;White et al 2014).…”

“…Error bars indicate means Ϯ SE. Additional procedural details of our electrophysiology approach and technical details of our equipment and measurements can be found in White et al (2014).…”

“…Even though the basal properties of simple spikes are determined intrinsically, sensory-induced afferent activity can influence their ultimate firing characteristics (Armstrong and Rawson 1979;Thach 1967;Wise et al 2010). They are characterized by a relatively regular pattern and high frequency of firing (ϳ40 -50 Hz) (Ruigrok 2011;Shin et al 2007), although they can fire over a wide range of frequencies in both anesthetized (Bosman et al 2010;de Solages et al 2008;White et al 2014) and awake preparations (Chaumont et al 2013;Cheron et al 2009;Goossens et al 2004;Shin et al 2007;Witter et al 2013;Zhou et al 2014). The second type of spike is called the "complex" spike.…”

WGA‐Alexa Conjugates for Axonal Tracing

“…Here, zebrinII was used at 1:250 (kind gift from Dr. Richard Hawkes; R. Hawkes, University of Calgary; Alberta; Canada Cat# ZebrinII Lot# RRID:AB_2315622; Figure 4). Please see our previous publications for additional details on our specific procedures for immunohistochemistry (Sillitoe and Hawkes, 2002; Sillitoe et al, 2010; White et al, 2014; White et al, 2016a; White et al, 2016b). …”