Neurons must establish and stabilize connections made with diverse targets, each with distinct demands and functional characteristics. At Drosophila neuromuscular junctions (NMJs), synaptic strength remains stable in a manipulation that simultaneously induces hypo-innervation on one target and hyper-innervation on the other. However, the expression mechanisms that achieve this exquisite target-specific homeostatic control remain enigmatic. Here, we identify the distinct target-specific homeostatic expression mechanisms. On the hypo-innervated target, an increase in postsynaptic glutamate receptor (GluR) abundance is sufficient to compensate for reduced innervation, without any apparent presynaptic adaptations. In contrast, a target-specific reduction in presynaptic neurotransmitter release probability is reflected by a decrease in active zone components restricted to terminals of hyper-innervated targets. Finally, loss of postsynaptic GluRs on one target induces a compartmentalized, homeostatic enhancement of presynaptic neurotransmitter release called presynaptic homeostatic potentiation (PHP) that can be precisely balanced with the adaptations required for both hypo- and hyper-innervation to maintain stable synaptic strength. Thus, distinct anterograde and retrograde signaling systems operate at pre- and post-synaptic compartments to enable target-specific, homeostatic control of neurotransmission.

Presynaptic homeostatic plasticity (PHP) adaptively enhances neurotransmitter release following diminished postsynaptic glutamate receptor (GluR) functionality to maintain synaptic strength. While much is known about PHP expression mechanisms, postsynaptic induction remains enigmatic. For over 20 years, diminished postsynaptic Ca2+ influx was hypothesized to reduce CaMKII activity and enable retrograde PHP signaling at the Drosophila neuromuscular junction. Here, we have interrogated inductive signaling and find that active CaMKII colocalizes with and requires the GluRIIA receptor subunit. Next, we generated Ca2+-impermeable GluRs to reveal that both CaMKII activity and PHP induction are Ca2+-insensitive. Rather, a GluRIIA C-tail domain is necessary and sufficient to recruit active CaMKII. Finally, chimeric receptors demonstrate that the GluRIIA tail constitutively occludes retrograde homeostatic signaling by stabilizing active CaMKII. Thus, the physical loss of the GluRIIA tail is sensed, rather than reduced Ca2+, to enable retrograde PHP signaling, highlighting a unique, Ca2+-independent control mechanism for CaMKII in gating homeostatic plasticity.

Ionotropic glutamate receptors (GluRs) are targets for modulation in Hebbian and homeostatic synaptic plasticity and are remodeled by development, experience, and disease. Although much is known about activity-dependent mechanisms that regulate GluR composition and abundance, the role of glutamate itself in these processes is unclear. To determine how glutamate sculpts GluR receptive fields, we have manipulated synaptically released glutamate and generated precise CRISPR mutations in the two postsynaptic GluR subtypes at the Drosophila neuromuscular junction, GluRA and GluRB. We first demonstrate that GluRA and GluRB compete to establish postsynaptic receptive fields, and that proper GluR abundance and localization can be orchestrated in the absence of any synaptic glutamate release. However, excess glutamate release adaptively tunes postsynaptic GluR abundance, echoing GluR receptor scaling observed in mammalian systems. Unexpectedly, when GluRA vs GluRB competition is eliminated, excess glutamate homeostatically regulates GluRA abundance, while GluRB abundance is now insensitive to glutamate modulation. Finally, Ca2+ impermeable GluRA receptors are no longer sensitive to homeostatic regulation by glutamate. Thus, excess glutamate, GluR competition, and Ca2+ signaling collaborate to selectively target GluR subtypes for homeostatic regulation at postsynaptic compartments.

27Neurons must establish and stabilize connections made with diverse targets, each with distinct 28 demands and functional characteristics. At Drosophila neuromuscular junctions, synaptic 29 strength remains stable in a manipulation that simultaneously induces hypo-innervation on one 30 target and hyper-innervation on the other. However, the expression mechanisms that achieve 31 this exquisite target-specific homeostatic control remain enigmatic. Here, we identify the distinct 32 target-specific homeostatic expression mechanisms. On the hypo-innervated target, an increase 33 in postsynaptic glutamate receptor (GluR) abundance is sufficient to compensate for reduced 34 innervation, without any apparent presynaptic adaptations. In contrast, a target-specific 35 reduction in presynaptic neurotransmitter release probability is reflected by a decrease in active 36 zone components restricted to terminals of hyper-innervated targets. Finally, loss of 37 postsynaptic GluRs on one target induces a compartmentalized, homeostatic enhancement of 38 presynaptic neurotransmitter release called presynaptic homeostatic potentiation that can be 39 precisely balanced with the adaptations required for both hypo-and hyper-innervation to 40 maintain stable synaptic strength. Thus, distinct anterograde and retrograde signaling systems 41 operate at pre-and post-synaptic compartments to enable target-specific, homeostatic control of 42 neurotransmission. 43 44 45 46 47 48 49 50 51 52 3 INTRODUCTION 53Synapses are spectacularly diverse in their morphology, physiology, and functional 54 characteristics. These differences are reflected in the molecular composition and abundance of 55 synaptic components at heterogenous synaptic subtypes in central and peripheral nervous 56 systems (Atwood and Karunanithi, 2002; Branco and Staras, 2009; O'Rourke et al., 2012). 57Interestingly, the structure and function of synapses can also vary substantially across terminals 58 of an individual neuron (Fekete et al., 2019; Grillo et al., 2018; Guerrero et al., 2005) and drive 59 input-specific presynaptic plasticity (Letellier et al., 2019). Both Hebbian and homeostatic 60 plasticity mechanisms can work locally and globally at specific synapses to tune synapse 61 function, enabling stable yet flexible ranges of synaptic strength (Diering and Huganir, 2018; 62 Turrigiano, 2012; Vitureira and Goda, 2013). For example, homeostatic receptor scaling globally 63 adjusts GluR abundance, subtype, and/or functionality at dendrites (Turrigiano and Nelson, 642004) yet there is also evidence for synapse specificity (Béïque et al., 2011; Hou et al., 2008; 65 Sutton et al., 2006). Although a number of studies have begun to elucidate the factors that 66 enable both local and global modes of synaptic plasticity at synaptic compartments, it is less 67 appreciated how and why specific synapses undergo plasticity within the context and needs of 68 information transfer in a neural circuit. 69One major force that sculpts the heterogeneity of synaptic strength is impose...