Homer1 Scaffold Proteins Govern Ca2+ Dynamics in Normal and Reactive Astrocytes

Abstract

In astrocytes, the intracellular calcium (Ca2+) signaling mediated by activation of metabotropic glutamate receptor 5 (mGlu5) is crucially involved in the modulation of many aspects of brain physiology, including gliotransmission. Here, we find that the mGlu5-mediated Ca2+ signaling leading to release of glutamate is governed by mGlu5 interaction with Homer1 scaffolding proteins. We show that the long splice variants Homer1b/c are expressed in astrocytic processes, where they cluster with mGlu5 at sites displaying intense local Ca2+ activity. We show that the structural and functional significance of the Homer1b/c-mGlu5 interaction is to relocate endoplasmic reticulum (ER) to the proximity of the plasma membrane and to optimize Ca2+ signaling and glutamate release. We also show that in reactive astrocytes the short dominant-negative splice variant Homer1a is upregulated. Homer1a, by precluding the mGlu5-ER interaction decreases the intensity of Ca2+ signaling thus limiting the intensity and the duration of glutamate release by astrocytes. Hindering upregulation of Homer1a with a local injection of short interfering RNA in vivo restores mGlu5-mediated Ca2+ signaling and glutamate release and sensitizes astrocytes to apoptosis. We propose that Homer1a may represent one of the cellular mechanisms by which inflammatory astrocytic reactions are beneficial for limiting brain injury.

Methamphetamine Addiction Vulnerability: The Glutamate, the Bad, and the Ugly

“…The mechanism(s) through which Homer2 maintains NAC glutamate are still unknown (27). Recently, however, mGlu5-Homer1 interactions were identified as important in exocytotic glutamate release from astrocytes (46). This raises the intriguing possibility that Homer2 regulation of calcium dynamics in astrocytes may be involved in the capacity of this isoform to regulate extracellular glutamate levels in forebrain.…”

Regulation and Function of Activity-Dependent Homer in Synaptic Plasticity

“…These observations corroborate closely with those from neocortex pyramidal neurons [95], both illustrating how activity-induced Homer isoforms permit a rise in intracellular calcium concentration. In astrocytes, however, the interruption of Homer1b/c-mediated mGluR5-ER interactions by Homer1a leads to a decrease in calcium signaling and glutamate release [97]. …”

Insights into the release mechanism of astrocytic glutamate evoking in neurons NMDA receptor‐mediated slow depolarizing inward currents

“…Efflux of endogenous glutamate from hippocampal slices was measured by a Glutamate Assay kit (Sigma-Aldrich) according to the manufacturer's instructions and as previously described (Buscemi et al, 2017). Coronal slices of 250 μm thin were obtained with a Thermo Scientific vibratome (Microm HM 650 V).…”

“…Slices were incubated in ACSF at least for 1 hr at 34 before processing. Efflux of endogenous glutamate from hippocampal slices was measured by a Glutamate Assay kit (Sigma-Aldrich) according to the manufacturer's instructions and as previously described (Buscemi et al, 2017). Briefly, for individual glutamate measurements, two to four hippocampal slices were lodged in a cuvette inside a bath at 37 C in a buffer solution containing (in mM): 120 NaCl, 3.1 KCl, 1.25 NaH 2 PO 4 , 25 Na-HEPES, 1 MgCl 2 , 4 glucose, 2 CaCl 2 , 0.001 TTX at pH 7.4 with 5% CO 2 , 95% O 2 .…”