A Comparison of Two Alternatively Spliced Forms of a Metabotropic Glutamate Receptor Coupled to Phosphoinositide Turnover

Abstract

A comparison of the pharmacological and physiological properties of the metabotropic glutamate 1 alpha and 1 beta receptors (mGluR1 alpha and mGluR1 beta) expressed in baby hamster kidney (BHK 570) cells was performed. The mGluR1 beta receptor is an alternatively spliced form of mGluR1 alpha with a modified carboxy terminus. Immunoblots of membranes from the two cell lines probed with receptor-specific antipeptide antibodies showed that mGluR1 alpha migrated with an M(r) = 154,000, whereas mGluR1 beta migrated with an M(r) = 96,000. Immunofluorescence imaging of receptors expressed in BHK 570 cells revealed that the mGluR1 alpha receptor was localized to patches along the plasmalemma and on intracellular membranes surrounding the nucleus, whereas mGluR1 beta was distributed diffusely throughout the cell. Agonist activation of the mGluR1 alpha and the mGluR1 beta receptors stimulated phosphoinositide hydrolysis. At both receptors, glutamate, quisqualate, and ibotenate were full agonists, whereas trans-(+)-1-aminocyclopentane-1,3-dicarboxylate appeared to act as a partial agonist. The stimulation of phosphoinositide hydrolysis by mGluR1 alpha showed pertussis toxin-sensitive and insensitive components, whereas the mGluR1 beta response displayed only the toxin-insensitive component. The mGluR1 alpha and mGluR1 beta receptors also increased intracellular calcium levels by inducing release from intracellular stores. These results indicate that the different carboxy terminal sequences of the two receptors directly influences G protein coupling and subcellular deposition of the receptor polypeptides and suggest that the two receptors may subserve different roles in the nervous system.

Modulatory effects of NMDA on phosphoinositide responses evoked by the metabotropic glutamate receptor agonist 1S,3R‐ACPD in neonatal rat cerebral cortex

“…The EC50 for this response (16.6IM) is in good agreement with values previously obtained in guinea-pig cerebral cortex (Jones & Roberts, 1993) and primary rat cerebro-cortical cultures (Birrell & Marcoux, 1993), although the magnitude of the response reported here is much greater. If it is assumed that the phosphoinositide-generating activity of trans-ACPD (a mixture of 1S,3R and lR,3S enantiomers) resides in the 1S,3R enantiomer (Irving et al, 1990;Jones & Roberts, 1993), then comparable EC50 values for trans-ACPD-stimulated phosphoinositide turnover have been obtained for mGlu-R5 (50 fM; Abe et al, 1992) and mGluRlb and MGluRlP (37 and 106 jLM respectively ;Pickering et al, 1993) in cells transfected to express specific metabotropic receptor subtypes.…”

“…accumulation can be almost precisely mimicked, including the apparent shift to the left of the concentration-response curve, by increasing the extracellular K+ concentration from 4.7 to 30 mm (data not shown). Thus, although mGluR coupling to PI-PLC is mediated, at least in part, via a pertussis toxinsensitive G-protein (Abe et al, 1992;Aramori & Nakanishi, 1992;Pickering et al, 1993), the PI-PLC(s) activated by this transduction pathway appears to exhibit similar Ca2+-sensitivity to PI-PLC(s) activated by receptors linked via pertussis toxin-insensitive G-proteins. ure 2) and Ins(1,4,5)P3 mass accumulation was lower than that observed in the presence of lS,3R-ACPD only (see Figure 6).…”

“…Of these, glutamate receptor agonists have been shown to stimulate phosphoinositide turnover in cells expressing mGluRla/,B and mGluR5 (Abe et al, 1992;Tanabe et al, 1992;Pickering et al, 1993). The mGluRs linked to phosphoinositide turnover exhibit pharmacological differences (Abe et al, 1992;Aramori & Nakanishi, 1992;Tanabe et al, 1992;Pickering et al, 1993), differential susceptibility to pertussis toxin (Abe et al, 1992;Aramori & Nakanishi, 1992;Pickering et al, 1993) and have been shown to be differentially expressed in rat brain (Abe et al, 1992;Martin et al, 1992;Fotuhi et al, 1993).…”

“…Since the original cloning and functional expression of a glutamate receptor (mGluRl) belonging to the guanine nucleotide binding protein-coupled superfamily (Masu et al, 1991), a number of structurally-related receptors have been described (Abe et al, 1992;Tanabe et al, 1992;Pickering et al, 1993). Of these, glutamate receptor agonists have been shown to stimulate phosphoinositide turnover in cells expressing mGluRla/,B and mGluR5 (Abe et al, 1992;Tanabe et al, 1992;Pickering et al, 1993).…”

Rapid agonist mediated phosphorylation of the metabotropic glutamate receptor 1α by protein kinase C in permanently transfected BHK cells

“…This cell line has been extensively characterized in the literature with respect to expression of the receptor, its agonist selectivity and signal transduction pathway [15,24,25]. The antibody A4, raised against an intracellular sequence (EFVYER-EGNTEEDEL) from mGluR I c~ has also been extensively characterised [26][27][28].…”