Program of Neural and Behavioral Sciences, Department of Physiology and Pharmacology, State University of New York Downstate Medical Center, Brooklyn, New York 11203
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ABSTRACT |
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Pancreatic islets contain
ionotropic glutamate receptors that can modulate hormone secretion. The
purpose of this study was to determine whether islets express
functional group III metabotropic glutamate (mGlu) receptors. RT-PCR
analysis showed that rat islets express the mGlu8 receptor subtype.
mGlu8 receptor immunoreactivity was primarily displayed by
glucagon-secreting -cells and intrapancreatic neurons. By
demonstrating the immunoreactivities of both glutamate and the
vesicular glutamate transporter 2 (VGLUT2) in these cells, we
established that
-cells express a glutamatergic phenotype. VGLUT2
was concentrated in the secretory granules of islet cells, suggesting
that glutamate might play a role in the regulation of glucagon
processing. The expression of mGlu8 by glutamatergic cells also
suggests that mGlu8 may function as an autoreceptor to regulate
glutamate release. Pancreatic group III mGlu receptors are functional
because mGlu8 receptor agonists inhibited glucagon release and
forskolin-induced accumulation of cAMP in isolated islets, and
(R,S)-cyclopropyl-4-phosphonophenylglycine, a group III mGlu receptor
antagonist, reduced these effects. Because excess glucagon secretion
causes postprandial hyperglycemia in patients with type 2 diabetes,
group III mGlu receptor agonists could be of value in the treatment of
these patients.
glutamate; glucagon; vesicular glutamate transporter 2; (R,S)-4-phosphonophenylglycine; (S)-3,4-dicarboxyphenylglycine; (R,S)-cyclopropyl-4-phosphonophenylglycine; cyclic adenosine monophosphate
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INTRODUCTION |
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GLUTAMATE, the major excitatory neurotransmitter in the mammalian central nervous system (CNS; Ref. 20), is also found in cells outside the CNS, including neurons in the enteric nervous system (19) and in endocrine cells in the pineal gland (21) and pancreatic islets (13). Enteric neurons appear to utilize glutamate as an excitatory transmitter (19). Pinealocytes use glutamate as either a paracrine- or autocrine-like chemical transmitter to inhibit melatonin synthesis (21). The role of glutamate in pancreatic islets is not yet fully understood.
Pancreatic -cells, which synthesize and secrete glucagon, express a
high-affinity, Na+-dependent glutamate/aspartate
transporter (30) and secrete glutamate through
Ca2+-dependent, regulated exocytosis (33).
Moreover, very recently, differentiation-associated
Na+-dependent inorganic phosphate (DNPI) cotransporter, a
vesicular glutamate transporter associated with synaptic vesicles in
central glutamatergic neurons (7, 8), was identified in
pancreatic
-cells (10), indicating that
-cells are
capable of both storing and secreting glutamate. These findings support
the possibility that pancreatic
-cells utilize glutamate as an
intercellular signaling molecule.
Pancreatic islets express both
N-methyl-D-aspartate (NMDA) and non-NMDA
[(RS)--amino-3-hydroxy-5-methyl-4-isoxazole propionic acid
(AMPA-type); kainate] ionotropic glutamate receptors, and activation
of AMPA receptors can potentiate insulin and glucagon secretion by
modulating the intrinsic properties of
- and
-cells (3, 13,
17, 31, 32). The endocrine pancreas might also be influenced by
glutamate via activation of metabotropic glutamate (mGlu) receptors.
The mGlu receptors are G protein-coupled receptors that are divided into three groups on the basis of sequence homology, pharmacology, and signal transduction mechanisms (for review, see Ref. 24). Group I (mGlu1, -5) receptors are coupled via Gq/G11 to phospholipase C. Both group II (mGlu2, -3) and group III (mGlu4, -6, -7, -8) receptors are coupled to Gi and inhibit stimulated cAMP formation when expressed in cell lines. In addition, members of the group II and group III mGlu receptors typically function as autoreceptors and inhibit the release of glutamate or other neurotransmitters when activated (24).
All mGlu receptor subtypes are expressed in the CNS, where they have been implicated in several aspects of physiology and pathology (5). However, mGlu receptors are also found in peripheral organs, as shown by the presence of mGlu4 receptors in taste buds (4), mGlu5 receptors in enteric neurons (18), and mGlu8 receptors in osteoblasts (11). Recently, a novel mGlu receptor-like protein was identified, and high levels of the receptor were found in the pancreas (6). In addition, islet cells appear to display mGlu2 and mGlu3 receptor immunoreactivity (14). This prompted us to search for the expression of mGlu receptors in the pancreatic islets.
We now report, for the first time, that group III mGlu receptors are
expressed in rat islets, that mGlu8 receptors are expressed by
glutamatergic -cells, and that activation of group III mGlu receptors inhibits glucagon release from isolated islets. These findings are consistent with the idea that islet mGlu8 receptors are
autoreceptors and support the possibility that
-cells utilize glutamate as an autocrine- and/or paracrine-like chemical transmitter, resulting in mGlu8 receptor-mediated inhibition of glucagon secretion.
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METHODS |
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Tissue. Adult female rats (Sprague-Dawley; 150-200 g) were euthanized by CO2 inhalation and then decapitated. The Animal Care and Use Committee of SUNY Downstate Medical Center approved this procedure. The pancreas was removed and washed with oxygenated (95% O2-5% CO2) Krebs solution of the following composition (mM): 121 NaCl, 5.9 KCl, 2.5 CaCl2, 14.3 NaHCO3, 1.3 NaH2PO4, 1.2 MgCl2, and 12.7 glucose.
RNA isolation and RT-PCR.
Total RNA (5 µg) from rat pancreas, freshly isolated islets (see
Isolation of islets), and brain, prepared using the TRIzol reagent (Ref. 15; Life Technologies, Gaithersburg, MD),
was reverse transcribed at 42°C (1 h) with the use of random primers and murine leukemia virus reverse transcriptase (Applied
Biosystems). This served as a template for PCR using
Taq DNA polymerase. PCR was performed using the primer
sequences as listed in Table 1. Primer
sequences were similar to those published previously (12). We used glyceraldehyde-3-phosphate dehydrogenase (GAPDH) as a housekeeping gene. After an initial denaturation step at 94°C for 5 min, the PCR conditions were as follows: 95°C for 15 s, 52°C
for GAPDH and 50°C for mGlu receptors for 1 min, and 72°C for 2 min
for 35 cycles, followed by a final extension step at 72°C for 7 min.
In controls, reverse transcriptase was omitted. The results of this
amplification were used to ensure successful mRNA isolation without
genomic DNA contamination. The PCR products were resolved in 2%
agarose gel with ethidium bromide. The PCR products were subcloned into
EcoRI-digested pGEM-T vector (Promega) for sequencing via
dye termination cycle sequencing (ABI Pyramid Automated Sequencer,
Perkin Elmer). The best match of the sequences was determined using the
gapped BLAST and PSI-BLAST programs.
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Immunocytochemistry. The pancreas was fixed for 24 h with 4% paraformaldehyde in 0.1 M phosphate buffer (pH 7.4). After fixation, the tissue was washed in phosphate-buffered saline (PBS), cryoprotected overnight (at 4°C) in PBS containing 30% sucrose, embedded in OCT (TissueTek), frozen with liquid N2, and sectioned (10 µm) using a cryostat-microtome (Leica). To locate mGlu8 receptor protein in the tissue by immunocytochemistry, preparations were incubated with 4% normal horse serum with Triton X-100 (0.5%) in PBS for 30 min. The preparations were then exposed overnight to guinea pig polyclonal antibodies generated against a peptide corresponding to amino acids 894-908 of rat and human mGlu8 receptor (diluted 1:1,000; Chemicon International, Temecula, CA; Ref. 26). After a wash with PBS, the preparations were incubated with donkey anti-guinea pig secondary antibodies coupled to Rhodamine Red-X (RRX; Jackson ImmunoResearch Laboratories, West Grove, PA) or fluorescein isothiocyanate (FITC; Jackson), diluted 1:500, for 3 h. The preparations were washed again with PBS, and then the tissues were coverslipped with Vectashiel (Vector Laboratories, Burlingame, CA).
Double-label immunocytochemistry was used to identify the cells that display mGlu8 receptor-like immunoreactivity. Double labeling was made possible by using primary antibodies raised in different species in conjunction with species-specific secondary antibodies [donkey anti-rat, donkey anti-mouse, donkey anti-guinea pig (Kirkegaard and Perry, Gaithersburg, MD), donkey anti-goat (Jackson ImmunoResearch Labs, diluted 1:500)] coupled to contrasting fluorophores (FITC or RRX, as in Immunocytology). Primary antibodies were against glucagon (guinea pig polyclonal, diluted 1:500, Linco Research), glutamate (mouse monoclonal, diluted 1:500, Incstar), insulin (mouse monoclonal, diluted 1:2,000, Chemicon International), pancreatic polypeptide (PP; affinity-purified rabbit polyclonal, diluted 1:1,000), somatostatin (affinity-purified rabbit polyclonal, diluted 1:1,000, Incstar), synaptophysin (mouse monoclonal, diluted 1:200, Sigma), or DNPI (rabbit polyclonal, diluted 1:1,500), which has been renamed, vesicular glutamate transporter 2 (VGLUT2; Ref. 7). The antibody to VGLUT2 was raised to a bacterial fusion protein containing the cytoplasmic COOH terminus of the rat protein. By Western blot analysis, the antibody recognizes VGLUT2, but not vesicular glutamate transporter 1 (VGLUT1), stably expressed in PC12 cells (7). Control sections, which were used to determine the level of nonspecific staining, included incubating sections without primary antibody and/or blocking the primary antibody by preincubation (24 h) with the corresponding peptide (10-20 µg/ml) before incubation of the antibody with the tissue. In both cases, no specific staining was observed. Preparations were examined by using a Radiance 2000 laser scanning confocal microscope (Bio-Rad, San Fransisco, CA) attached to an Axioskop 2 microscope (Zeiss). Usually, 5-10 optical sections were taken at 1.0-µm intervals. Images of 512 × 512 pixels were obtained and processed using Adobe Photoshop 6.0 (Adobe Systems, Mountain View, CA) and printed using a Tektronix Phaser 440 printer.Western blot analysis. The Ready Western Single Tissue Blot from rat pancreas was purchased from DNA Technologies (Gaithersburg, MD). Seventy-five micrograms of protein were applied to the lane, and the blot was blocked (30 min) with 5% nonfat dry milk in washing buffer (0.5% Tween 20 in Tris-buffered saline) and then probed with affinity-purified primary antibody against mGlu8 (1:2,000) for 2 h. After a wash, the Vectastain ABC kit for guinea pig IgG (Vector Laboratories) was used to detect the bound antibody, and the blot was visualized by using the Vector SG substrate kit (Vector Laboratories).
Electron microscopy. Postembedding staining with colloidal gold was carried out as described by Fremeau et al. (7). In general, isolated islets (see Isolation of islets) were fixed with 4.0% paraformaldehyde and 0.1% glutaraldehyde in 0.1 M sodium phosphate buffer (pH 7.4) at room temperature for 3 h and embedded in L.R.White (Electron Microscopy Sciences). Ultrathin sections, mounted on Formvar-coated grids, were then etched on drops of fresh 1% H2O2 (30 min) and washed in distilled H2O. Sections were incubated with the primary antiserum (rabbit anti-VGLUT2, diluted 1:500 for 4 days at 4°C) followed by several washes and then incubated with 6-nm particles of colloidal gold (diluted 1:100) adsorbed to goat anti-rabbit IgG Fab-fragments (Electron Microscopy Sciences) for 90 min at room temperature. After a rinse, the sections were contrasted with uranyl acetate and lead citrate and observed in a JEOL JEM-100C electron microscope. Control sections were incubated without primary antibody.
Isolation of islets. Islets were isolated from the rat pancreas as previously described (16). In brief, after cannulation of the common bile duct, the pancreas was injected with 15 ml of Hanks' solution (in mM: 137 NaCl, 5.36 KCl, 4.17 NaHCO3, 0.34 Na2HPO4, 0.44 KH2PO4, 0.81 MgSO4 · H2O, 1.26 CaCl2 · 2H2O, 2.8 D-glucose) containing 8 mg/ml collagenase P (Boehringer Mannheim, Mannheim, Germany) through the intrapancreatic duct. The pancreas was removed and digested for 15 min at 37°C in a shaking water bath followed by dilution and washing with Hanks' solution containing 0.5% BSA. Individual islets were handpicked and placed into culture dishes.
Glucagon secretion assay.
Glucagon release was measured at 37°C after static incubations.
Groups of eight size-matched islets were preincubated for 30 min in 500 µl extracellular (oxygenated Krebs) solution and 2.8 mM
D-glucose. The preincubation medium was aspirated
and discarded. The islets were then resuspended in 200 µl of
extracellular solution in the absence or presence of the specific group
III mGlu receptor agonist (R,S)-4-phosphonophenylglycine [(R,S)PPG;
Ref. 9]. When the effect of the group III mGlu receptor
antagonist (RS)--cyclopropyl-4-phosphonophenylglycine (CPPG; Ref.
29) was to be investigated, this compound was added to the
preincubation medium before the addition of the agonist. At the end of
the test incubation (1 h), the medium was aspirated and assayed for
glucagon by use of a rat glucagon enzyme-linked immunosorbent assay kit
(Peninsula Laboratory, San Carlos, CA) with human glucagon as standard.
cAMP assay.
Rat pancreatic islets were isolated as in Isolation of
islets and cultured overnight in RPMI medium 1640 containing 10%
fetal bovine serum and antibiotics. The islets were washed thoroughly with Krebs solution before use. After 30 min of the preincubation described, 15 size-matched islets were incubated for 30 min at 37°C
with 0.5 ml of Krebs solution containing 2.8 mM D-glucose, 5 mg/ml BSA, and 1 mM 3-isobutyl-1-methylxanthine (IBMX) and various substances as indicated. At the end of the incubation, islets were
transferred into 250 µl of 50 mM sodium acetate with 20 mM EGTA and
homogenized. Islet extracts were spun at 13,000 g for 20 min, and the supernatant was retained and stored at 20°C until assayed. The total cAMP content of islets was determined as picomoles cAMP per milliliter by using a cAMP-enzyme immunoassay kit from Assay
Designs (Ann Arbor, MI).
Chemicals and drugs. (R,S)PPG, CPPG, and (S)-3,4-dicarboxyphenylglycine (DCPG) were obtained from Tocris Cookson (Ballwin, MO). All other chemicals were purchased from Fisher Scientific (Fair Lawn, NJ) or Sigma Chemical (St. Louis, MO).
Statistical analysis. Data are expressed as means ± SE, with differences between groups determined by Student's t-test.
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RESULTS |
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Group III mGlu receptor expression in the rat pancreas.
Group III mGlu receptor expression in the pancreas was analyzed
utilizing RT-PCR. As expected, the brain was positive for mGlu4 (Fig.
1B, lane 1), mGlu6
(Fig. 1B, lane 2), mGlu7 (Fig. 1B,
lane 3), and mGlu8 expression (Fig. 1A,
lane 2), with expected bands at 340, 363, 321, and 440 bp,
respectively. The pancreas (Fig. 1C) and isolated islets
(Fig. 1D) were also positive for mGlu8 receptor expression
(Fig. 1, C and D, lane 5), whereas no expression
was detected in these tissues for mGlu4 (Fig. 1, C and
D, lane 2), mGlu6 (Fig. 1, C and D, lane 3),
or mGlu7 (Fig. 1, C and D, lane 4)
receptors. RT-PCR analysis was confirmed after we subcloned and
sequenced the PCR products. These results suggest that the pancreas,
and specifically the islets, express only the gene encoding mGlu8
receptors, whereas the brain expresses the genes encoding all four
group III mGlu receptor subtypes.
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Localization of mGlu8 receptor immunoreactivity in the pancreas. If mGlu8 receptors were present in the pancreas, then, as in other areas where these receptors exist, the pancreas would be expected to contain sites that can be demonstrated with mGlu8 receptor-selective antibodies (12). In both the pancreatic islets (Fig. 1E) and ganglia (Fig. 1F), a population of cells displayed mGlu8 receptor immunoreactivity. In general, immunolabeling was punctate and localized to cell bodies. In addition, mGlu8 receptor immunoreactivity was found on numerous nerve fibers (Fig. 1F), suggesting that mGlu8 receptors are located presynaptically. No immunoreactivity was found in control sections that were processed without the primary antiserum (not illustrated). Moreover, Western blotting with the antibody against mGlu8 showed one labeled band of protein in rat pancreas membrane (Fig. 1G). The labeled protein had an apparent molecular mass of 100 kDa, which is in agreement with the molecular mass deduced from the cDNA sequence of mGlu8.
Identification of mGlu8 receptor-immunoreactive islet cells.
Further studies were done to chemically identify the islet cells with
mGlu8 receptor immunoreactivity. Three nonoverlapping groups of islet
cells were identified. One group displayed glucagon immunoreactivity
(Fig. 2, A-C) and 100% of
glucagon-immunoreactive -cells contained mGlu8 receptor. The other
groups contained insulin (Fig. 2, D-F) or pancreatic
polypeptide immunoreactivity (Fig. 2, G-I), and subsets of
these cells expressed mGlu8. In contrast, mGlu8 receptor
immunoreactivity was not found in somatostatin-containing cells (Fig.
2, J-L).
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mGlu8 receptor immunoreactivity is displayed by islet cells expressing a glutamatergic phenotype. In the CNS, several studies have shown that group III mGlu receptors are expressed by glutamatergic neurons and function as autoreceptors that regulate the physiological release of glutamate (24). This prompted us to determine whether mGlu8-immunoreactive islet cells express a glutamatergic phenotype.
mGlu8 receptor immunoreactivity was displayed by islet cells that contained glutamate (Fig. 3, A-C) and synaptophysin (not illustrated), a marker for synaptic-like vesicles. Moreover, these cells expressed VGLUT2 (Fig. 3, D-F), which was found in all glutamate-immunoreactive islet cells (Fig. 3, G-I). In addition, glutamate/VGLUT2-immunoreactive cells contained glucagon (Fig. 3, J-L), suggesting that mGlu8 is expressed by glutamatergic
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VGLUT2 is associated with secretory granules in pancreatic islet cells. To elucidate the role of glutamate in pancreatic islets, we localized VGLUT2 in isolated islets employing postembedding electron microscopic immunocytochemistry with immunogold. We focused on the localization of VGLUT2 in cells situated near the border of the islets, because the majority of VGLUT2-positive cells were found in this region.
Analysis at the electron microscopic level confirmed the presence of high levels of VGLUT2 in a subset of islet cells. Cells were identified as glucagon-producing
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Activation of mGlu8 receptors inhibits glucagon release and cAMP
production.
To assess whether the expressed mGlu8 receptors in pancreatic -cells
cells are functional, we determined the effects of the group
III-specific agonist (R,S)PPG (9) on glucagon release and
cAMP production in isolated islets.
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DISCUSSION |
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The present study demonstrates, for the first time, that group III mGlu receptors are found in pancreatic islets. With the use of RT-PCR analysis, rat islets were shown to express the gene encoding mGlu8. Expression of mGlu8 was corroborated by immunocytochemical data localizing mGlu8 receptor protein in the islet cells. Moreover, functional studies demonstrated that islet group III mGlu receptors might play a role in modulating pancreatic endocrine secretion.
mGlu8 receptors were primarily expressed by -cells, although a
subset of insulin- and PP-secreting cells also appeared to display
mGlu8 receptor immunoreactivity. In addition to islets, mGlu8 receptor
immunoreactivity was also found on neural elements within the pancreas.
Pancreatic nerve cell bodies and fibers displayed intense mGlu8
receptor immunoreactivity; therefore, group III mGlu receptors may be
involved in modulating synaptic transmission in pancreatic ganglia. The
presence of mGlu8 in all
-cells suggested that activation of group
III mGlu receptors might affect glucagon release.
Application of the group III mGlu receptor agonist (R,S)PPG
significantly reduced glucagon secretion in isolated islets, and the
effect of (R,S)PPG was inhibited by the group III mGlu receptor antagonist CPPG. Thus islet mGlu8 receptors appear to be functional and
negatively modulate the release of glucagon when activated. The
inhibitory effect of mGlu8 receptors on glucagon release could be, at
least in part, attributed to its interfering with the adenylate cyclase/cAMP system, since both (R,S)PPG and DCPG significantly inhibited forskolin-stimulated cAMP production in isolated islets. In
addition, the effects of agonists were suppressed by CPPG. Taken
together, these findings suggest that glutamate, through the activation
of mGlu8 receptors located on the -cells, inhibits adenylate
cyclase, thereby decreasing glucagon secretion.
To date, very few selective group III mGlu receptor agonists have been identified, and only (R,S)PPG and DCPG have been reported to have any degree of selectivity for mGlu8 (9, 28). Nevertheless, despite the lack of selective ligands, it seems likely that mGlu8 is involved, because islets only express the gene encoding mGlu8 receptors and both (R,S)PPG and DCPG are highly potent in this system. The potency of (R,S)PPG and DCPG to produce a response in isolated islets appears to be greater than in mGlu8 receptor-expressing cells (28, 29). An increase in the efficiency of an agonist to elicit an effect is commonly attributed to an increase in the receptor reserve for the agonist (23). Whether there is a large receptor reserve at pancreatic mGlu8 receptors has not yet been determined; however, understanding the mechanisms associated with the supersensitivity of mGlu8 receptors in the islets could be relevant to the pathophysiology of diabetes.
Islet cells that expressed mGlu8 receptors displayed a glutamatergic phenotype. mGlu8 receptor-positive cells contained both glutamate and the vesicular glutamate transporter VGLUT2, which were found co-localized in the same cells. VGLUT2 is one of two types of vesicular glutamate transporter expressed in central glutamatergic neurons (7, 8), the other being brain-specific Na+-dependent inorganic phosphate cotransporter, renamed VGLUT1 (Refs. 1 and 27). Because the VGLUT accumulates glutamate into synaptic vesicles (1, 7, 8, 27), VGLUT is a selective marker of glutamatergic cells.
Glutamate is released from TC6 cells, clonal mouse pancreatic
-cells, through a Ca2+-dependent exocytotic mechanism
(33). Immunoelectron microscopy revealed a large number of
synaptophysin-positive clear vesicles in these cells. From these
findings, it was concluded that
TC6 cells, like central
glutamatergic neurons (1, 7, 8, 27) and pinealocytes
(10), store glutamate in synaptophysin-containing vesicles
(33). In support of this hypothesis,
TC6 cells, as well
as
-cells in rat islets, have recently been found to express VGLUT2
(10). However, whether VGLUT2 is located in
synaptophysin-containing vesicles was not determined.
In the present study, we confirmed that -cells in rat islets express
VGLUT2. Interestingly, VGLUT1 immunoreactivity is also displayed by a
subset of pancreatic
-cells (Q. Tong and A. L. Kirchgessner,
unpublished observations). Therefore,
-cells are probably capable of
storing and secreting glutamate. The presence of punctate VGLUT2
immunoreactivity in the cytoplasm of these cells suggested that VGLUT2
might localize to vesicles. Labeling with gold particles in isolated
islets showed that VGLUT2 was localized in the secretory granules of a
subset of islet cells, presumably the glucagon-producing
-cells,
because they contained an abundance of round granules of high density
and a narrow halo. In addition, the majority of VGLUT2-reacting gold
particles were found along or near the secretory granule membrane,
although some diffusion of gold particles did occur. This finding
supports the idea that VGLUT2 transports glutamate into secretory
granules and that glutamate might play a role in the regulation of
glucagon processing. Whether glutamate is localized in secretory
granules in islet cells is not yet known; however,
D-aspartate is stored in secretory granules in
pheochromocytoma PC12 cells (22).
Although the physiological role of glutamate in the islets is not fully
understood, glutamate appears to regulate the secretion of islet
hormones by way of its binding to both ionotropic (3, 13, 17, 31,
32) and mGlu receptors. Glutamate acting via ionotropic
receptors exerts an excitatory action and stimulates both insulin and
glucagon secretion (13, 31). In this study, we showed that
glutamate acting via the group III mGlu8 receptor exerts an inhibitory
action on glucagon release. Moreover, mGlu8-positive islet cells
display a glutamatergic phenotype. In the CNS, the mGlu8 receptor is
known to couple to Gi and function as an autoreceptor to
negatively modulate the release of glutamate when activated (24). As in the CNS, group III mGlu receptors in -cells
may also provide negative feedback to limit further release of
glutamate under normal (and/or pathological) conditions of glutamate
release. In this way, glutamate may act as an autocrine- and/or
paracrine-like chemical transmitter, resulting in inhibition of
glucagon synthesis. Because lack of suppression of glucagon secretion
can cause postprandial hyperglycemia in subjects with type 2 diabetes
(25), group III mGlu receptor agonists could be of value
in the treatment of these patients.
In summary, the results from this study have shown that 1)
pancreatic islets express the gene encoding mGlu8; 2) mGlu8
receptor immunoreactivity is displayed primarily by -cells, which
display a glutamatergic phenotype; and 3) activation of
group III mGlu receptors inhibits glucagon release and
forskolin-stimulated cAMP production in isolated islets. Expression of
mGlu8 in
-cells that contain glutamate and VGLUT2 suggests that this
subtype may act as an autoreceptor to inhibit glutamate release. The
presence of both ionotropic and mGlu receptors in the islets allows
glutamate to play complex roles in endocrine secretion.
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ACKNOWLEDGEMENTS |
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We thank Drs. Robert H. Edwards and Robert T. Fremeau for providing the antibody to VGLUT2, Drs. Jeffrey Conn and Min-tsai Liu for helpful discussions, and Wei Quan for assistance with electron microscopy.
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FOOTNOTES |
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This work was supported by National Institutes of Health Grant NS-35951 and The American Diabetes Association (to A. L. Kirchgessner).
Address for reprint requests and other correspondence: A. Kirchgessner, Dept. of Physiology and Pharmacology, Box 29, SUNY Downstate Medical Center, 450 Clarkson Ave., Brooklyn, NY 11203 (E-mail: akirchgessner{at}netmail.hscbklyn.edu).
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
First published February 5, 2002;10.1152/ajpendo.00460.2001
Received 15 October 2001; accepted in final form 30 January 2002.
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