Multilayer control of KaiR1D-autoreceptor function by the auxiliary protein Neto

Kainate-type glutamate receptors and their dedicated auxiliary protein Neto function at both pre- and postsynaptic sites to regulate the activity of synaptic networks. However, attributing specific synaptic functions to Neto and/or kainate receptors is challenging. Here we focus on Drosophila KaiR1D receptors, which modulate synaptic transmission at neuromuscular junction, and elucidate the role of Neto in the regulation of autoreceptor activities and neurotransmitter release. We show that Neto-α limits the presynaptic accumulation and function of KaiR1D autoreceptors in vivo. Using outside-out patch recordings, we demonstrate that Neto-α modulates the KaiR1D gating properties, slowing desensitization and attenuating the block by intracellular polyamines and extracellular toxins. Neto-α also promotes the axonal distribution of KaiR1D, but this function is not critical for the KaiR1D-dependent regulation of synaptic transmission. Instead, Neto-α increases the charge transfer and likely the KaiR1D-mediated influx of Ca²⁺ in the presynaptic compartment leading to increased neurotransmitter release. Our data demonstrate that Neto-α provides multiple layers of modulation to KaiR1D autoreceptors to ensure proper neurotransmitter release. These findings broaden our view on how auxiliary subunits shape channel gating and subcellular distribution, suggesting that coordinated regulation of receptor function and localization represents an ancestral strategy to safeguard synaptic stability.