Our previous work delineated a novel interaction between the CaSR and a class of cellular adhesion proteins known as the integrins (Tharmalingam et al., 2011). Identification of the integrins as CaSR interacting proteins was initially determined in rat thyroid carcinoma cells from CaSR immuno-affinity pull-down experiments combined with mass spectrometry. Pharmacological stimulation of the CaSR using the positive allosteric modulator NPS R-568 potently enhanced thyroid carcinoma cell adhesion and migration. Evidence that these effects occurred via the integrins was indicated by the demonstration that the enhancement of cell-ECM adhesion by NPS R-568 could be largely blocked by the addition an integrin blocking peptide (GRGDSP) that competes with fibronectin for β1-integrin binding. Additional confirmation was obtained whereby shRNA-mediated knock down of β1-containing integrins reduced CaSR-mediated cell adhesion and migration. We concluded that (a) the CaSR is associated with β1-containing integrins in a macromolecular protein complex and (b) that stimulation of the CaSR promotes cell adhesion and migration via integrin activation (Tharmalingam et al., 2011).

The integrins are a family of proteins that mediate cell-to-cell contact by binding to the ECM. Integrins are cell adhesion molecules that interact with the ECM to provide a dynamic link between the extracellular adhesion molecules and the intracellular actin cytoskeleton. These proteins are unique in that they are able to bind to ECM proteins and induce intracellular signaling cascades, and are also capable of conveying intracellular signaling messages to the ECM by modulating their binding affinity for the ECM. By mediating cell-ECM interactions integrins play an essential role in cellular migration. They are also involved in other cellular processes including cellular differentiation, proliferation, embryogenesis, CNS development, inflammation and cancer (Iwamoto and Calderwood, 2015; Maartens and Brown, 2015; Wang et al., 2015).

Integrins are heterodimers of type 1 membrane-spanning glycoproteins composed of an α and a β subunit (Figure 1). In mammals, there are 18 α and 8 β subunits that form 24 heterodimeric pairs (see Campbell and Humphries, 2011 and Maartens and Brown, 2015 for reviews). Both α and β integrin subunits are characterized by a very large extracellular domain of 700–1100 residues, a single transmembrane domain, and a short cytoplasmic domain of 30–50 residues, with both subunits held together via non-covalent interactions. The outermost N-terminal region of the α integrin subunit that faces the extracellular side of the cell is composed of a head structure with seven-bladed β-propellers, which connects to the leg structure composed of a thigh, calf-1 and calf-2 domains (Barczyk et al., 2010). A region between the thigh and calf-1 domain known as the “α subunit genu” acts as a pivot point for integrin domain extension.

In the resting state integrins adopt a conformation where the integrin leg structures are severely bent, generating a closed topology where the head domain are juxtaposed to the leg regions close to the cell membrane (Figure 1). In this conformation the integrin ligand binding sites are not optimal for binding ECM proteins, and therefore the bent conformation represents a low-affinity state. The presence of ECM ligands induces an intermediate-affinity conformation in which a switch-blade like extension of the head region shifts away from the legs (Luo and Springer, 2006; Barczyk et al., 2010). Destabilization of the α/β integrin association fully activates the integrins into a high-affinity conformation (O'Toole et al., 1991; Lu et al., 2001), and transmembrane domain separation is necessary for integrin signaling (Wang and Luo, 2010).

From the intracellular side, integrins do not bind actin directly, but rather indirectly by proteins they recruit with their cytoplasmic tails. The dynamic complex of proteins at cell–ECM adhesions has been termed the integrin adhesome. A crucial adhesome component is the cytoskeletal-associated protein talin, which can simultaneously bind the cytoplasmic tail of integrin β subunits and actin, as well as coordinate the solicitation of other proteins to the adhesome (Critchley, 2009). Integrin inside-out signaling is mediated by the binding of talin to the β integrin C-terminal tail, a process that is controlled by the small GTPase Rap1 and its effectors such as RapL and RIAM (Mehrbod et al., 2013). Rap1 is activated by various GPCRs that are activated by chemoattractant stimulation. Stimulation of membrane spanning receptors mediates cell adhesion and migration by activating signaling cascades that lead to integrin inside-out signaling (Mehrbod et al., 2013). Upon separation of the integrin α and β subunits, the integrins adopt a high affinity active conformation that allows binding of ECM proteins. Ligand (ECM) occupancy and integrin clustering in-turn activates intracellular signaling cascades that mediate cell cytoskeletal changes, a process known as integrin outside-in signaling (Figure 1).

Upon binding to ECM proteins, integrin dimers cluster together laterally on the membrane via cell surface insertion of integrins localized in caveolae-rich microdomains (Maheshwari et al., 2000). Integrin clustering induces actin filament assembly and reorganization of actin filaments into larger stress fibers in turn causes further integrin clustering, providing a positive feedback loop that maintains enhanced matrix binding. The aggregation of ECM proteins, integrins, cytoskeletal proteins, and signaling kinases forms unique structures known as focal adhesion complexes (Parsons et al., 2010). In particular, integrin ligation and clustering activates focal adhesion kinase (FAK), which in-turn activates Src-family kinases (Mitra and Schlaepfer, 2006; Hu and Luo, 2013). Activated Src phosphorylates cytoskeletal proteins such as paxillin, actin bundling protein α-actinin and vinculin to form nascent adhesion structures. Binding of these initial cytoskeletal proteins recruits further actin nucleating proteins that result in focal adhesion complex and actin stress-fiber formation. These cytoskeletal changes mediated by integrin outside-in signaling ultimately allow cells to adhere and spread over the ECM proteins.

Intriguingly, and also similar to the CaSR, the integrins contain multiple divalent cation binding sites in the extracellular domain (Xiong et al., 2002). The integrin cation binding sites can be occupied by Ca²⁺ or by Mn²⁺ ions. However, the integrins differ from the CaSR in that the cations are necessary but not sufficient for integrins to convert from the inactive bent conformation into the active extended conformation. Both the presence of cations bound to the multiple cation binding sites is required, along with the direct physical association with ECM ligands for integrins to attain the extended structure and concomitant activation (Dai et al., 2015). Based on these features, we hypothesize that a rise in extracellular Ca²⁺ ions would serve to both prime the integrin heterodimer and activate the CaSR (see further discussion below).

Integrins are crucial players in several aspects of bone tissue biology (see Marie et al., 2014 for review). Several integrin heterodimers including α5β1 integrin, α2β1 integrin, and α4β1 integrin have been shown to play important roles in bone formation and maintenance, and in bone cell differentiation (Hamidouche et al., 2009; Guan et al., 2012; Marie, 2012). In general, activation of integrins on osteoblasts leads to cell differentiation. In bone (and other tissues) ECM–integrin binding induces the recruitment and phosphorylation of focal adhesion kinase and activation of phosphatidylinositol 3-kinase (PI3K) and mitogen-activated protein kinase (MAPK) ERK1/2, leading to activation of the transcription factor RUNX2 and osteoblast differentiation.

Of special interest here in light of the identification of the CaSR- β1 integrin complexes, is that β1-containing integrins play an important role in osteoblast differentiation. Direct evidence supporting this concept comes from mice expressing a dominant-negative β1 integrin subunit in mature osteoblasts. These mutant mice show reduced bone mass, defective bone formation and loss of ICAP-1, an important modulator of β1 integrin activation resulting in defective osteoblast differentiation and delayed bone formation (Globus et al., 2005; Brunner et al., 2011). The a5β1 integrin heterodimer which binds to the ECM protein fibronectin, is essential for osteoblast formation, and α5 integrin suppression reduces osteoblast differentiation, whereas its overexpression facilitates osteoblast differentiation in cultured mesenchymal skeletal cells. These effects are mediated via FAK/MAPK ERK1/2 signaling (Hamidouche et al., 2009). It has also been shown that integrin αvβ3 is critical for CYR61-mediated BMP-2 expression and subsequent osteoblastic differentiation through activation of the ILK/MAPK ERK1/2 signaling pathway in osteoblasts (Su et al., 2010). Similarly, α2β1, a major receptor for collagen type 1, was reported to control collagen synthesis in osteoblasts in mice (Stange et al., 2013). This integrin heterodimer plays a role in mesenchymal skeletal cells osteogenic differentiation and survival through activation of ROCK, FAK, and MAPK ERK1/2 signaling. Together, these findings provide insight into the role of integrin-mediated signaling pathways in the control of osteoblast differentiation, fate, and function (Marie et al., 2014).

Integrins have been intensively studied in the context of cancer with hundreds of literature citations on the topic; therefore, we will restrict our discussion here to the most pertinent findings on the association of β1-containing integrins and metastasis to bone.

β1 integrins are activated in metastatic prostate cancer cells and mediate increased prostate cancer cell metastasis to lymph nodes and bone. Dissemination of prostate cancer cells to the bone marrow occurs early in the disease. Cancer cell proliferation and active metastases after a period of undetectable disease is known as tumor dormancy. Dormancy-reactivation has been studied in patient-derived xenograft lines where the propensity to proliferate through tumor cell contact with each other, and with bone marrow stroma was demonstrated (Ruppender et al., 2015). Proliferating prostate cancer cells displayed tumor cell-cell contact and integrin clustering after ECM contact (e.g., fibronectin) in bone marrow stroma. Activation of myosin light chain kinase, a downstream effector of integrin-β1 and TGF- β2, in non-proliferating cells promoted cell proliferation. This activity leads to the deactivation of growth suppressor like E2F4 and activation of cell cycle regulator CDK6 thereby promoting cell proliferation (Ruppender et al., 2015). These data illustrate the involvement of integrins in the escape of prostate cancer cells from dormancy.

Similar to CaSR expression in metastatic prostate cancers, β1-containing integrins have been shown to be constitutively activated in prostate cancer cell lines with high metastatic potential, but not in cancer cell lines with low metastatic potential (Lee et al., 2013). Activation of β1 integrins promotes resistance to anoikis. Anoikis is a form of programmed cell death that is induced by anchorage-dependent cells detaching from the surrounding ECM. Usually cells stay close to the tissue to which they belong since the communication between proximal cells as well as between cells and ECM provide essential signals for growth or survival. When cells are detached from the ECM, there is a loss of normal cell–matrix interactions, and they may undergo anoikis. However, metastatic tumor cells may escape from anoikis and invade other organs. Resistance to anoikis occurs via stimulation of FAK/Akt-mediated survival pathway and neutralizing β1 integrin activation using an anti-β1 integrin antibody reduced prostate cancer metastasis in vivo (Lee et al., 2013). In addition, talins which are adaptor proteins that regulate focal adhesion signaling by conjugating integrins to the cytoskeleton, bind integrins and are essential for integrin activation. In this context, talin has been shown to be important for β1 integrin activation, and Cdk5-mediated phosphorylation of talin leading to β1 integrin activation was shown to increase the metastatic potential of prostate cancer cells (Jin et al., 2015).

Integrins are also involved in breast cancer metastasis. Bone metastasis affects more than 70% of advanced breast cancer patients, but the underlying molecular mechanisms remain unclear. Although some evidence suggests that integrins mediate adhesion of malignant cells to bone ECM, their role during bone colonization is also not fully understood. The role of β1 integrins in bone colonization has been investigated in vitro and in vivo in tissue-engineered humanized bone models (Thibaudeau et al., 2015). In vitro, bone- metastatic breast cancer cells with suppressed integrin β1 expression showed reduced attachment, spreading, and migration within human bone matrix compared to control cells. Cell proliferation in vitro was not affected by β1 integrin knockdown, but tumor growth in vivo within humanized bone microenvironments was significantly inhibited upon β1 integrin suppression. Tumor cells invaded bone marrow spaces in the humanized bone and formed osteolytic lesions whereas osteoclastic bone resorption was, however, not reduced by β1 integrin knockdown (Thibaudeau et al., 2015). These observations indicate that β1-containing integrins likely play a prominent role in bone colonization.

Some data are also available linking integrins and bone metastasis in renal carcinomas. Renal carcinoma cells that form bone metastasis constitutively activate osteoclasts, which then induce bone resorption and release pro-proliferative factors. As noted above, this phenomenon also occurs in bone metastasis of other types of tumors including breast and prostate cancer. Constitutive activation of osteoclasts leads to the release of fibronectin and collagen which can facilitate cancer cell attachment via integrins to ECM and vascular endothelial cells. Cell adhesion of bone metastasizing cells to fibronectin and to collagen I (but not adhesion to collagen IV) is high and bone has a high concentration of these two ECM proteins (Ode et al., 2010). In one study, 9 renal cell carcinoma primary cell lines and 30 renal cell carcinoma tissue specimens (normal and tumor tissue) collected from 3 patients with no metastasis, and 10 samples with lung or bone metastasis within 5 years after nephrectomy were collected and analyzed (Haber et al., 2015). Compared to renal cell carcinoma cells from patients without metastasis, the migration of cells from patients with bone metastasis was enhanced 14-fold, and adhesion to fibronectin and collagen I were both enhanced 6-fold. Notably, proliferation was decreased in metastasizing cells. In addition, elevated activity of Akt and FAK and integrin α5 expression, and reduced PTEN expression was observed in primary cells from patients with bone metastasis compared to non-metastasizing cells. Thus, (Haber et al., 2015) concluded that increased expression of integrin α5 and downstream signaling via Akt could help tumor cells to adhere to ECM proteins and facilitate migration to bone tissue.

In summary, three lines of evidence lend support to the hypothesis that CaSR-integrin complexes may be a factor in some forms of cancer metastasis: (1) the findings outlined above and others indicating a role for β1-containing integrins in cancer metastasis to bone, (2) the considerable body of evidence demonstrating elevated CaSR expression in prostate, breast, and kidney cancers that metastasize to bone, and (3) the presence of CaSR-β1 integrin macromolecular complexes in thyroid carcinoma cells (Tharmalingam et al., 2011). We postulate that in circulating tumor cells, CaSR-integrin protein complexes could function together as a cell surface detection mechanism for attaching to the ECM components of the bone microenvironment (Figure 2). The CaSR could act as the calcium sensor and become activated by the (presumed) high extracellular Ca²⁺; in bone tissue this would be followed in turn by activation of the associated integrins on the tumor cell to signal attachment to bone ECM proteins such as fibronectin and collagen. It should be emphasized that this proposed sequence of events is speculative and further work is needed to rigorously test the idea.

CaSR expression in the central nervous system (CNS) was first demonstrated using a rat striatal cDNA library where it was determined that the receptor is expressed in both neurons and glia (Ruat et al., 1995). In adults, the CaSR is localized to nerve terminals with discrete punctate expression. Upon depolarization, the synaptic space contains elevated Ca²⁺. Due it is localization within nerve terminals, the CaSR may regulate neurotransmitter disposition in response to synaptic Ca²⁺ changes (Ruat et al., 1995; Ruat and Traiffort, 2013). In the immature brain the receptor is expressed in several regions including the orbital cortex, cerebellum (see below), and hippocampus (Ferry et al., 2000; Tharmalingam et al., 2016). Analysis of rat hippocampal mRNA and protein expression showed that peak CaSR expression occurs from postnatal day 12 to day 24, followed by a 3-fold decrease to adult levels (Chattopadhyay et al., 1997). Cellular analysis demonstrated CaSR expression in pyramidal cells of all the layers of hippocampus and in granule cells of the dentate gyrus.

Studies on of CaSR knockout mice during postnatal brain development have indicated that the CaSR plays an important role in the differentiation of numerous brain cell types. For example, CaSR null mice exhibited developmental delay due to impaired differentiation as assessed by reduced expression of proliferating cell nuclear antigen, neuronal specific nuclear protein (neuronal differentiation marker), glial fibrillary acidic protein (glial differentiation marker), and myelin basic protein (oligodendrocyte differentiation marker) when compared to age-matched wild-type littermates (Liu et al., 2013). Brain weight and size were drastically reduced in the CaSR null mice and cell proliferation was also decreased in the CaSR knockout animals. Concurrent deletion of the CaSR with parathyroid hormone gene corrected for the effects of hyperparathyroidism, hypercalcemia, hypophosphatemia, and whole-body growth retardation (Liu et al., 2013). However, further analysis of these animals established that brain cell proliferation was normal whereas differentiation was greatly reduced in the CaSR-PTH double knockout animals. The role of the CaSR in differentiation vs. proliferation was also investigated using cultured neural stem cells derived from the subventricular zones of CaSR null neonatal mice, which exhibited normal proliferation capacity but decreased differentiation capacity when compared with wild-type animals (Liu et al., 2013). Taken together, the CaSR is crucial for differentiation of neurons, glia and oligodendrocytes. Delayed brain development in CaSR null newborn mice is therefore the result of direct effects of the CaSR on cell differentiation, and indirect CaSR effects on cell proliferation via dysregulation of parathyroid hormone secretion and serum Ca²⁺ (Liu et al., 2013).

The CaSR expression pattern in adult rodent brain differs substantially from the immature brain. The regions of moderate to high expression during development do not express the receptor, or express it at very low levels, in the adult rodent brain. However, the CaSR is expressed in a few regions of the adult CNS including the olfactory bulbs, area postrema, and the subfornical organ (Ferry et al., 2000; Yano et al., 2004). The subfornical organ is a hypothalamic thirst center; its neurons project onto supraoptic and paraventricular nuclei where they control the activity of vasopressin-secreting neurons (Ruat and Traiffort, 2013). Expression in the subfornical organ suggests that the receptor is also involved in central fluid and electrolyte balance (Yano et al., 2004). The adult cerebral arteries also display an intense network of CaSR immunoreactive fibers demonstrating possible functional role in vessel innervation. Studies on adult CaSR neuronal expression have determined that the CaSR controls neuronal excitability by regulating ion channel function (Phillips et al., 2008; Vyleta and Smith, 2011). These findings reinforce the idea that the CaSR modulates Ca²⁺-dependent neuronal functions in both the immature and in the adult brain.

β1 integrins are expressed in all cell types in the CNS and integrin heterodimers containing the β1 integrin are indispensable for many functions in the brain where the β1 subunit forms obligate heterodimers with numerous α integrin subunits to create functional receptor complexes. Each brain region and cell type expresses its own repertoire of α integrin subunits resulting in different integrin receptor expression profile and function (reviewed by Pinkstaff et al., 1999). In the developing nervous system, spatial and temporal changes in integrin expression parallel changes in neurogenesis, differentiation, and migration (Schmid and Anton, 2003). In early CNS development, neuronal migration during establishment of the cerebral and cerebellar cortex requires β1 integrin-mediated adhesion and migration (Hatten, 1999). Neurons originating from the ventricular zone migrate radially using glial cells and detach from the glial fibers at the cortical plate to establish connections at their final resting position. Loss of β1 integrins in neurons results in grossly disorganized structures in the most superficial layer of the cortex (Milner and Campbell, 2002), and selective deletion of β1 integrins in glia resulted in failure of glial cell contact formation with the meningeal basement membrane and blocked neuronal radial migration (Blaess et al., 2004; Frick et al., 2012). Therefore, normal assembly of the cerebral and cerebellar cortex requires β1 integrin mediated neuronal migration and glial adhesion (Milner and Campbell, 2002).

The well-established migrational patterns of immature cerebellar granule-cell precursor neurons (GCPs) in the early postnatal cerebellum provides an excellent model system to delineate the molecular properties of neuronal migration. GCPs originate in the rhombic lip during late embryonic stages and then migrate rostrally to a secondary germinal layer, known as the external granule-cell layer of the cerebellum, where they continue to proliferate after birth. In rodents GCPs proliferate in the periphery of the external granule layer-cerebellar basement membrane interface over postnatal days 5–20 (Figure 3). Various factors such as laminin (expressed on the basement membrane), sonic hedge-hog (released from Purkinje cells), brain-derived neurotropic factor (released from Bergmann glia), and insulin-like growth factors contribute to GCP proliferation. GCPs that have proliferated and exited the cell-cycle undergo tangential migration within the external granule layer (Figure 3). Tangentially migrating GCPs attach to the Bergmann glial processes and undergo radial migration whereby the GCPs migrate radially along the Bergmann glia fibers through the cerebellar molecular layer and the Purkinje cell layer, and into their final resting position in the internal granule-cell layer (Chedotal, 2010). By 3 weeks after birth GCP proliferation ceases and the remaining GCPs complete migration into the internal granule layer, which is retained in the mature brain while the external granule layer disappears. In the adult CNS cerebellar granule cells are the most numerous class of neurons in the vertebrate brain.

The role of β1 integrins in cerebellar GCP proliferation and migration has been well documented. Potent stimulants of GCP proliferation such as sonic hedgehog and SDF-1 have been shown to induce GCP proliferation only in the presence of β1 integrin mediated adhesion to laminin (Klein et al., 2001; Blaess et al., 2004). β1 integrins control GCP migration by binding to ECM proteins such as laminin, vitronectin, reelin, etc., which aid in the homing of GCPs from the external granule layer to the internal granule layer (Pons et al., 2001; Borghesani et al., 2002; Porcionatto, 2006). Thus, GCP migration relies heavily on integrin activation. Notably, defects in GCP migration result in medulloblastoma, the most common cause of malignant brain tumors in young children (see below).

We have recently demonstrated that the CaSR, together with integrins, functions as a cell-surface chemoattractant receptor expressed on the surface of cerebellar GCPs to facilitate GCP differentiation into mature granule cells, and to simultaneously guide their movement during cerebellar maturation (Tharmalingam et al., 2016). A robust but transient increase in CaSR expression in GPCs was observed that peaked during the second postnatal week, the period of maximal GPC migration. Our results support a model whereby the CaSR acts as a chemoattractant sensor to detect a gradient of extracellular Ca²⁺, while integrin heterodimers in conjunction with ECM proteins function as the driving force to mediate cell movement (Tharmalingam et al., 2016). The data also indicated that CaSR-mediated stimulation of ERK2 and Akt phosphorylation provides a biochemical link between Ca²⁺-sensing and the integrin-mediated movement of GCPs (Figure 5; also see further discussion below).

A key question to be addressed is, what is the source of the CaSR stimulus in the immature cerebellum? One possibility is that activation of the CaSR may be mediated by local extracellular Ca²⁺ released by the Bergmann glia. Tangentially migrating unipolar GCPs in the external granule layer sprout a third axon that allows the GCPs to attach to Bergmann glial fiber tracts to undergo radial migration into the internal granule layer. Bergmann glial fibers have been shown to generate radially expanding transglial Ca²⁺ waves that initiate at a central point and expand radially to encompass multiple Bergmann glial processes (Hoogland et al., 2009). The Ca²⁺ waves generated by Bergmann glia could locally activate the CaSR expressed on GCPs to promote radial migration. Another potential source of CaSR activation may be from polyamines. Polyamines such as spermine and spermidine are potent agonists of the CaSR and polyamine concentrations are elevated during postnatal cerebellar development and then subsequently decline to adult levels (Jasper et al., 1982). The period of increased polyamines corresponds to the peak period of CaSR expression and GCP migration. Moreover, the initial step in the synthesis of the polyamines is mediated by the enzyme ornithine decarboxylase. In the cerebellum of rats injected with α-difluoromethylornithine, an inhibitor of ornithine decarboxylase, granule cells fail to migrate properly and the development of the cerebellar cortex is stalled in an immature state (Bartolome et al., 1985). Taken together these findings indicate that polyamines might act alone, or in conjunction with Ca²⁺ ions, to stimulate the CaSR and guide GCP migration in the developing brain.

Defects in GCP migration result in medulloblastoma, the most common cause of malignant brain tumors in children under four (Hatten and Roussel, 2011; Remke et al., 2013; Macdonald et al., 2014). Medulloblastoma is a highly heterogeneous group of tumors characterized by aberrant GCP proliferation and migration. Unlike most neuronal populations, GCPs remain mitotically active after birth (Roussel and Hatten, 2011). Impaired migration causes GCPs to remain longer in the proliferative-niche of the external granule layer where they are more prone to transformational insults due to increased exposure to proliferation-inducing molecules. There are currently no effective treatments for medulloblastoma. Although a role for the CaSR in medulloblastoma has not been established, the identification of key proteins regulating GCP migration and progression is a major goal in medulloblastoma research, and could lead to novel approaches for the development of effective therapeutics.

Medulloblastoma has been modeled in vitro using DAOY cells which are transformed human cerebellar granule cells. Wei et al. (2015) studied the function of the CaSR in these cells and compared them to normal cerebellar granule cells in culture (Wei et al., 2015). They examined the CaSR in conjunction with OGR1 (also known as GPR68), an extracellular proton-sensing GPCR coupled to Gq and the release intracellular Ca²⁺. In addition to protons, this receptor is also activated by the benzodiazepine anxiolytic drug lorazepam (Huang et al., 2015). Previous work had established that OGR1 activation in DAOY cells leads to complex intracellular Ca²⁺ signals and activation of the ERK signaling pathway, thereby providing a mechanistic explanation of how the acidic environment may influence transformed cell function and/or survival. This action is lost on differentiation, suggesting a link between OGR1 activity and proliferative behavior of the transformed neurons (Huang et al., 2009). Curiously, Ca²⁺ signaling through OGR1 and CaSR appears to be opposite in the two cell types. In normal granule cells, OGR1-dependent Ca²⁺ signaling is slow and small while CaSR-dependent intracellular Ca²⁺ signaling is fast and large, whereas the opposite occurs in DAOY cells (Wei et al., 2015). It was suggested that the lack of inhibition of OGR1 signaling by CaSR in DAOY cells is likely due, at least in part, to low expression of functional CaSR in these cells, given that knockdown of CaSR in granule cells also leads to disinhibition of OGR1 signaling. Moreover, CaSR-dependent inhibition of OGR1 activity is absent in DAOY cells and intracellular acidification, which may accompany extracellular acidosis, inhibited CaSR responses but potentiated OGR1 responses (Wei et al., 2015).

OGR1 is functionally expressed in brain tumor cells and changes in its activity may be relevant to acidosis in tumors. Both OGR1 and CaSR have been implicated in several brain disorders that are exacerbated by changes in the concentrations of extracellular H⁺ and Ca²⁺ (Kingsley et al., 2007). The CaSR and OGR1 are also co-expressed in other tissues that experience extracellular acidification under physiological conditions including kidney, bone, and lung. Therefore, in addition to cerebellar neurons, a perturbation in the balance between OGR1 and CaSR signaling may contribute to the development and progression of other pathological conditions.

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.