Abnormal larval neuromuscular junction morphology and physiology in Drosophila Prickle isoform mutants with defective axonal transport and adult seizure behavior

Abstract

Previous studies have demonstrated that mutations of the Drosophila planar cell polarity gene prickle (pk) result in altered microtubule-mediated vesicular transport in larval motor axons, as well as adult neuronal circuit hyperexcitability and epileptic behavior. It is also known that mutant alleles of the prickle-prickle (pkpk) and prickle-spiny-legs (pksple) isoforms differ in phenotype but display isoform counterbalancing effects in heteroallelic pkpk/pksple flies to ameliorate adult motor circuit and behavioral hyperexcitability. We have further investigated the larval neuromuscular junction (NMJ) and uncovered robust phenotypes in both pkpk and pksple alleles (heretofore referred to as pk and sple alleles, respectively), including synaptic terminal overgrowth, as well as irregular motor axon terminal excitability, poor vesicle release synchronicity, and altered efficacy of synaptic transmission. We observed significant increase in whole-cell excitatory junctional potential (EJP) in pk homozygotes, which was restored to near WT level in pk/sple heterozygotes. We further examined motor terminal excitability sustained by presynaptic Ca2+ channels, under the condition of pharmacological blockade of Na+ and K+ channel function. Such manipulation revealed extreme Ca2+ channel-dependent nerve terminal excitability in both pk and sple mutants. However, when combined in pk/sple heterozygotes, such terminal hyper-excitability was restored to nearly normal. Focal recording from individual synaptic boutons revealed asynchronous vesicle release in both pk and sple homozygotes, which nevertheless persisted in pk/sple heterozygotes without indications of isoform counter-balancing effects. Similarly, the overgrowth at NMJs was not compensated in pk/sple heterozygotes, exhibiting an extremity comparable to that in pk and sple homozygotes. Our observations uncovered differential roles of the pk and sple isoforms and their distinct interactions in the various structural and functional aspects of the larval NMJ and adult neural circuits.