Biochemical and Biophysical Research Communications
Neuronal overexpression of IP3 receptor 2 is detrimental in mutant SOD1 mice
Highlights
► Overexpression of IP3R2 increases cytosolic Ca2+ concentrations evoked by bradykinin. ► Thy1.2–IP3R2 mice have increased expression of IP3R2 in the spinal cord and brain. ► Neuronal overexpression of IP3R2 shortens the lifespan of the ALS mice.
Introduction
A tight control of intracellular Ca2+ concentration is crucial for cell survival [1]. Motor neurons are especially vulnerable to disturbances in Ca2+ dynamics as they combine a low concentration of Ca2+ binding proteins with a relatively high amount of Ca2+-permeable AMPA-type glutamate receptors [2]. This could contribute to the selective vulnerability of the motor neurons during Amyotrophic Lateral Sclerosis (ALS). ALS is a progressive neurodegenerative disease, characterized by the selective loss of motor neurons and the denervation of muscle fibers, resulting in muscle weakness and paralysis. In Europe, the disease has an annual incidence of 2.7 cases per 100,000 people [3] and the disease duration post diagnosis is 3–5 years. In 10% of patients, ALS is a familial disease and 20% of these familial ALS patients contain mutations in the gene encoding superoxide dismutase 1 (SOD1). As the disease progression is indistinguishable between familial and sporadic ALS, common disease mechanisms are predicted. One of these mechanisms is excitotoxicity that causes neuronal death by overstimulation of the glutamate receptors [2]. The only effective treatment in ALS, riluzole, reduces the glutamatergic input [2]. In general, excitotoxicity induces an increased intracellular Ca2+ concentration, which is hazardous to the motor neurons. In addition, release of Ca2+ from the endoplasmic reticulum (ER) may further increase the intracellular Ca2+ concentration resulting in increased excitotoxicity [4].
In this study, we investigate the role of elevated expression of an ER Ca2+ release channel, inositol 1,4,5-trisphosphate receptor 2 (IP3R2) and concomitant increased IP3-induced ER Ca2+ release in ALS. To this end, we generated a mouse with a neuronal overexpression of IP3R2 and crossbred these mice with a mouse model for ALS, in which excitotoxicity contributes to motor neuron death [2].
Section snippets
Animal generation and housing
Mice overexpressing human SOD1G93A were purchased from The Jackson Laboratories (Bar Harbor, USA) and maintained on a C57BL/6 background. In order to create transgenic mice, murine itpr2 cDNA (8.1 kb) was cloned into the XhoI restriction site of the Thy1.2 expression cassette (kind gift from Novartis Pharma). A 15 kb DNA fragment was excised from the Thy1.2–IP3R2 vector with the restriction endonuclease Not1. The transgenic mice overexpressing itpr2 (Thy1.2–IP3R2) were created by pronuclear
IP3R2 overexpression increases agonist-induced Ca2+ signals in N2a cells
To investigate whether increased ER Ca2+ release through the IP3 receptor may be detrimental in ALS, we cloned the cDNA of the murine ITPR2 gene into the Thy1.2 expression cassette (Fig. 1A). The Thy1.2 expression cassette drives transgene expression starting at postnatal day 9 and especially in neurons [5]. IP3R2 is the most sensitive isoform of the receptor to IP3 [6], [7]. In the nervous system IP3R2 is mainly localized in astrocytes [8], [9], [10], [11], [12], [13]. The functional
Discussion
We studied the potential contribution of Ca2+ originating from intracellular stores and the role for ER Ca2+ release channels in neurons during ALS. A potential role of IP3R2 was previously suggested by genome wide association of single nucleotide polymorphisms (SNPs) in the ITPR2 gene with sporadic ALS [14], although this association was not found in other populations [15], [16]. Interestingly, an increase of IP3R2 gene expression is detected in blood samples of ALS patients [14]. Although IP3
Acknowledgments
This work was supported by grants from the “Fund for Scientific Research Flanders” (FWO-Vlaanderen, FWO G.0608.09N), the University of Leuven (KU Leuven, including GOA/11/014 and OTSTART1/10/044) and the Belgian Government (Interuniversity Attraction Poles, programme P7/16) of the Belgian Federal Science Policy Office. PVD holds a clinical investigatorship of FWO-Vlaanderen and WR is supported through the E. von Behring Chair for Neuromuscular and Neurodegenerative Disorders.
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2017, NeuroscienceCitation Excerpt :However, Sig1R agonists do not increase IP3 formation itself (Hayashi et al., 2000); rather, they prevent IP3R inhibition (Wu and Bowen, 2008). IP3 overexpression was shown to be detrimental both in mutant SOD1 mice (Staats et al., 2012a) and the N2 cell line expressing the same mutation (Watanabe et al., 2013). Furthermore, the genetic ablation of phospholipase C delta 1 was shown to prolong survival in G93A mice; phospholipase C delta 1 increases IP3, which releases Ca2+ from the ER via IP3Rs (Staats et al., 2013).
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2016, Advances in Biological RegulationCitation Excerpt :Inositol 1,4,5-triphosphate (IP3)-mediated Ca2+ efflux from the ER regulates various signaling pathways (Mikoshiba, 2015), and its defects have been reported to be involved in the neuronal toxicity. One of the inositol 1,4,5-triphosphate receptors (IP3R) subtypes, IP3R2 is upregulated in the blood samples from ALS patients and the neuronal overexpression of IP3R2 shortened the lifespan of SOD1 G93A mice (Van Es et al., 2007; Staats et al., 2012). However, the precise mechanism of disturbances in neuronal calcium homeostasis in the ALS context is still unclear.
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