Homer 2 tunes G protein–coupled receptors stimulus intensity by regulating RGS proteins and PLCβ GAP activities

Homers are scaffolding proteins that bind G protein–coupled receptors (GPCRs), inositol 1,4,5-triphosphate (IP3) receptors (IP3Rs), ryanodine receptors, and TRP channels. However, their role in Ca2+ signaling in vivo is not known. Characterization of Ca2+ signaling in pancreatic acinar cells from Homer2−/− and Homer3−/− mice showed that Homer 3 has no discernible role in Ca2+ signaling in these cells. In contrast, we found that Homer 2 tunes intensity of Ca2+ signaling by GPCRs to regulate the frequency of [Ca2+]i oscillations. Thus, deletion of Homer 2 increased stimulus intensity by increasing the potency for agonists acting on various GPCRs to activate PLCβ and evoke Ca2+ release and oscillations. This was not due to aberrant localization of IP3Rs in cellular microdomains or IP3R channel activity. Rather, deletion of Homer 2 reduced the effectiveness of exogenous regulators of G proteins signaling proteins (RGS) to inhibit Ca2+ signaling in vivo. Moreover, Homer 2 preferentially bound to PLCβ in pancreatic acini and brain extracts and stimulated GAP activity of RGS4 and of PLCβ in an in vitro reconstitution system, with minimal effect on PLCβ-mediated PIP2 hydrolysis. These findings describe a novel, unexpected function of Homer proteins, demonstrate that RGS proteins and PLCβ GAP activities are regulated functions, and provide a molecular mechanism for tuning signal intensity generated by GPCRs and, thus, the characteristics of [Ca2+]i oscillations

Homer binds TRPC family channels and is required for gating of TRPC1 by IP 3 receptors

Receptor signaling at the plasma membrane often releases calcium from intracellular stores. For example, inositol triphosphate (IP3) produced by receptor-coupled phospholipase C activates an intracellular store calcium channel, the IP3R. Conversely, stores can induce extracellular calcium to enter the cell through plasma membrane channels, too. How this reverse coupling works was unclear, but store IP3Rs were proposed to bind and regulate plasma membrane TRP cation channels. Here, we demonstrate that the adaptor protein, termed Homer, facilitates a physical association between TRPC1 and the IP3R that is required for the TRP channel to respond to signals. The TRPC1-Homer-IP3R complex is dynamic and its disassembly parallels TRPC1 channel activation. Homer's action depends on its ability to crosslink and is blocked by the dominant-negative immediate early gene form, H1a. Since H1a is transcriptionally regulated by cellular activity, this mechanism can affect both short and long-term regulation of TRPC1 function

Synaptic Inputs Compete during Rapid Formation of the Calyx of Held: A New Model System for Neural Development

Hallmark features of neural circuit development include early exuberant innervation followed by competition and pruning to mature innervation topography. Several neural systems, including the neuromuscular junction and climbing fiber innervation of Purkinje cells, are models to study neural development in part because they establish a recognizable endpoint of monoinnervation of their targets and because the presynaptic terminals are large and easily monitored. We demonstrate here that calyx of Held (CH) innervation of its target, which forms a key element of auditory brainstem binaural circuitry, exhibits all of these characteristics. To investigate CH development, we made the first application of serial block-face scanning electron microscopy to neural development with fine temporal resolution and thereby accomplished the first time series for 3D ultrastructural analysis of neural circuit formation. This approach revealed a growth spurt of added apposed surface area (ASA)>200 μm2/d centered on a single age at postnatal day 3 in mice and an initial rapid phase of growth and competition that resolved to monoinnervation in two-thirds of cells within 3 d. This rapid growth occurred in parallel with an increase in action potential threshold, which may mediate selection of the strongest input as the winning competitor. ASAs of competing inputs were segregated on the cell body surface. These data suggest mechanisms to select "winning" inputs by regional reinforcement of postsynaptic membrane to mediate size and strength of competing synaptic inputs

Synaptic Inputs Compete during Rapid Formation of the Calyx of Held: A New Model System for Neural Development

Hallmark features of neural circuit development include early exuberant innervation followed by competition and pruning to mature innervation topography. Several neural systems, including the neuromuscular junction and climbing fiber innervation of Purkinje cells, are models to study neural development in part because they establish a recognizable endpoint of monoinnervation of their targets and because the presynaptic terminals are large and easily monitored. We demonstrate here that calyx of Held (CH) innervation of its target, which forms a key element of auditory brainstem binaural circuitry, exhibits all of these characteristics. To investigate CH development, we made the first application of serial block-face scanning electron microscopy to neural development with fine temporal resolution and thereby accomplished the first time series for 3D ultrastructural analysis of neural circuit formation. This approach revealed a growth spurt of added apposed surface area (ASA) >200 μm(2)/d centered on a single age at postnatal day 3 in mice and an initial rapid phase of growth and competition that resolved to monoinnervation in two-thirds of cells within 3 d. This rapid growth occurred in parallel with an increase in action potential threshold, which may mediate selection of the strongest input as the winning competitor. ASAs of competing inputs were segregated on the cell body surface. These data suggest mechanisms to select “winning” inputs by regional reinforcement of postsynaptic membrane to mediate size and strength of competing synaptic inputs

Homeostatic scaling requires group I mGluR activation mediated by Homer1a

Homeostatic scaling is a non−Hebbian form of neural plasticity that maintains neuronal excitability and informational content of synaptic arrays in the face of changes of network activity. Here, we demonstrate that homeostatic scaling is dependent on group I metabotropic glutamate receptor activation that is mediated by the immediate early gene Homer1a. Homer1a is transiently upregulated during increases in network activity and evokes agonist−independent signaling of group I mGluRs that scales down the expression of synaptic AMPA receptors. Homer1a effects are dynamic and play a role in the induction of scaling. Similar to mGluR−LTD, Homer1a−dependent scaling involves a reduction of tyrosine phosphorylation of GluA2 (GluR2), but is distinct in that it exploits a unique signaling property of group I mGluR to confer cell−wide, agonist−independent activation of the receptor. These studies reveal an elegant interplay of mechanisms that underlie Hebbian and non−Hebbian plasticity