DrosophilaVoltage-Gated Sodium Channels Are Only Expressed in Active Neurons and Are Localized to Distal Axonal Initial Segment-like Domains

Abstract

In multipolar vertebrate neurons, action potentials (APs) initiate close to the soma, at the axonal initial segment. Invertebrate neurons are typically unipolar with dendrites integrating directly into the axon. Where APs are initiated in the axons of invertebrate neurons is unclear. Voltage-gated sodium (Na V ) channels are a functional hallmark of the axonal initial segment in vertebrates. We used an intronic Minos-Mediated Integration Cassette to determine the endogenous gene expression and subcellular localization of the sole Na V channel in both male and female Drosophila, para. Despite being the only Na V channel in the fly, we show that only 23 6 1% of neurons in the embryonic and larval CNS express para, while in the adult CNS para is broadly expressed. We generated a single-cell transcriptomic atlas of the whole third instar larval brain to identify para expressing neurons and show that it positively correlates with markers of differentiated, actively firing neurons. Therefore, only 23 6 1% of larval neurons may be capable of firing Na V -dependent APs. We then show that Para is enriched in an axonal segment, distal to the site of dendritic integration into the axon, which we named the distal axonal segment (DAS). The DAS is present in multiple neuron classes in both the third instar larval and adult CNS. Whole cell patch clamp electrophysiological recordings of adult CNS fly neurons are consistent with the interpretation that Na v -dependent APs originate in the DAS. Identification of the distal Na V localization in fly neurons will enable more accurate interpretation of electrophysiological recordings in invertebrates.

The connectome of the adult Drosophila mushroom body provides insights into function

“…The pedunculus consists of parallel KC axonal fibers that connect the CA and the lobes and is largely devoid of external innervation in the adult. Voltage-gated sodium channels are concentrated in the proximal peduculus where they are likely to serve as the initiation point for KC action potentials ( Ravenscroft et al, 2020 ). There are five MB lobes: α, β, α′, β′, and γ.…”

A connectome of theDrosophilacentral complex reveals network motifs suitable for flexible navigation and context-dependent action selection

“…This heterogeneity and compartmentalization make it challenging to evaluate the relative weight of different synaptic inputs to the neuron's synaptic output. Even for fly neurons that spike, action potential initiation sites are largely unknown (although see (Gouwens and Wilson, 2009;Ravenscroft et al, 2020)). Furthermore, spiking neurons may perform local circuit computations involving synaptic transmission without action potentials.…”

Compensatory variability in network parameters enhances memory performance in theDrosophilamushroom body

“…Therefore, an excitatory input would make a smaller contribution to a KC’s decision to spike the farther away it is from the spike initiation zone [50]. While the spike initiation zone cannot be directly observed in the connectome, the voltage-gated Na + channel para and other markers of the axon initial segment (also called the ‘distal axonal segment’) are concentrated at the posterior end of the peduncle, near where axons from KCs derived from the four neuroblast clones converge [51, 52]. This location can be approximated in the connectome as the posterior boundary of the ‘PED(R)’ region of interest (ROI) (magenta dots, Fig.…”

“…Note that calcium signals are often entirely restricted to one dendritic claw [26, 53]. Another caveat is that the posterior boundary of the peduncle is only an estimate (though a plausible one: [51, 52]) of the location of the spike initiation zone. However, inaccurate locations should only produce fictitious correlations for Fig.…”