the actions of glutamate , the main excitatory neuro-  transmitter in the mammalian central nervous system ( cns ) , can be finely modulated by metabotropic glutamate receptors ( mglurs ) [ 1 , 2 ] .
mglurs are g - protein coupled receptors and are divided into three groups based on sequence homology , pharmacological profile , and signal transduction mechanisms .
eight mglurs ( mglur1 - 8 ) have been identified and classified into three groups : group i , consisting of mglur1 and mglur5 , group ii , consisting of mglur2 and mglur3 , and group iii , consisting of mglur4 , mglur6 , mglur7 , and mglur8 .
group i receptors are coupled to phospholipase c ( plc ) activation , while group  ii and iii are associated with adenylate cyclase inhibition [ 3 , 4 ] .
mglurs modulate glutamatergic transmission at several levels depending on their expression at nerve terminals , postsynaptic sites , or glia [ 5 , 6 ] ( fig .
1 ) . group i mglurs  are mainly located at the postsynaptic regions , where  they increase neural excitability , whereas group ii and group iii are primarily located at presynaptic terminals and function as inhibitory auto- and hetero - receptors [ 7 , 8 ] . while mglur6 is exclusively expressed in the retina , the other mglurs are widely distributed throughout the nervous system .
group iii is the largest group of mglurs and the least well - characterized , likely due to the lack of selective pharmacologic agents .
selective allosteric ligands for group iii mglur subtypes were recently discovered ; this has made it possible to elucidate the role of each of these receptors in normal cns functioning and in models of neurologic and psychiatric disorders .
recently , a brain penetrant preferential agonist for mglur4 , ( 2s)-2-amino-4-(hydroxy(hydroxy(4-hydroxy-3-methoxy5nitrophenyl)methyl)phosphoryl)butanoic acid ( lsp1 - 2111 ) was identified .
preclinical studies suggest that lsp1 - 2111 has in vivo efficacy in models of parkinson s disease , anxiety , psychosis , fear learning , and memory .
these studies rationalize further investigations on the therapeutic benefits of mglur modulators that can finely tune glutamatergic transmission in an effort to treat various psychiatric and neurologic conditions .
out of the group iii mglurs , mglur7 is the most widely expressed throughout the cns [ 7 , 11 , 12 ] .
the highest density of mglur7 expression is in the olfactory bulb , hippocampus , hypothalamus , and sensory afferent pathways [ 11 , 13 - 15 ] .
mglur7 receptors are mainly located within the presynaptic active zone [ 11 , 14 , 16 ] where they serve as auto- or hetero - receptors , inhibiting glutamate or gaba release , respectively [ 16 , 17 ] ( fig .
1 ) . mglur7 receptor activation also leads to increased signaling pathways potentiating neurotransmitter release in cerebrocortical nerve terminal preparations from adult rats .
however , direct facilitation of neurotransmitter release by mglur7 has not yet been demonstrated on neurons in vitro or in vivo . in the globus pallidus and striatum ,
postsynaptic mglur7 receptors have also been observed on amacrine cells at retinal synapses , on olfactory bulb glomeruli , and on prefrontal cortex pyramidal neurons .
so far , five splice variants of the mglur7 receptor have been characterized , called mglur7a - e .
these splice variants exhibit different , but often overlapping , expression patterns [ 12 , 23 - 25 ] .
in addition to its cns localization , mglur7 is also expressed in peripheral tissues , such as the colon , stomach , and adrenal glands , and in hair cells and spiral ganglion cells of the inner ear .
mglur7 has low affinity for glutamate ; it is recruited only under high neurotransmitter concentrations and thus acts as an auto - receptor to inhibit further glutamate release .
l-2-amino-4-phosphonobutyrate ( l - ap4 ) can bind to mglur7 at high concentrations and inhibits glutamate release via n - type ca channel inhibition   ( fig .
recently , studies using the selective mglur7 positive allosteric agonist n , n'-bis(diphenylmethyl)-1,2-ethanediamine dihydrochloride ( amn082 ) and the mglur7 negative allosteric modulator 7-hydroxy-3-(4-iodophenoxy)-4h - chromen-4-one ( xap044 ) have demonstrated that mglur7 modulates excitatory / inhibitory transmission in the hippocampus [ 32 - 35 ] , thalamus , nucleus accumbens [ 38 , 39 ] , pag   and amygdala [ 41 - 43 ] .
specifically , amn082 decreases gaba and increases glutamate levels in the nucleus accumbens   and amygdala .
, mglur7 facilitates glutamatergic release , likely via interactions with the exocytose machinery . apart from gi / o - protein coupling and the consequent inhibition of adenylyl cyclase and camp formation
, mglur7 also inhibits n- and p / q - type ca channels in the transfected cerebellar granule cells , brainstem , and hippocampus [ 46 , 47 ] .
finally , mglur7 also modulate synaptic function through g - protein - coupled inwardly rectifying potassium channels ( girks ) .
mglur7 activity is finely modulated : calmodulin binds to the carboxyl terminus of mglur7 in a ca - dependent manner ( cam ) .
the cam - binding domain , located at the end of the seventh trans - membrane segment , can be competitively phosphorylated by protein kinase c ( pkc ) , which inhibits binding of the ca / cam complex to the receptor .
pkc inhibits mglur7 s effect on neurotransmitter release by preventing coupling of mglur7 receptors to their requisite g proteins .
pkc - mediated phosphorylation is also a key mechanism regulating constitutive and activity - dependent mglur7 expression .
two events , mglur7 phosphorylation by pkc and mglur7 binding to the pdz domain - containing protein pick1 , lead to increased mglur7 expression .
mglur7 expression is also increased by inhibitors of protein phosphatase 1 ( pp1 ) , which counteracts the action of pkc on mglur7 .
rodent studies suggest that the interaction of mglur7 and pick1 is critical to proper neural function , as disruption of the mglur7a - pick1 complex is sufficient to induce absence epilepsy - like seizures .
mglur7 modulates gabaergic interneuron synapse development through its interaction with elfn1 protein , whose abnormal expression during development is associated with epilepsy and attention deficit hyperactivity disorders .
mglur7 also prevents nmda - induced excitotoxicity of basal forebrain ( bf ) cholinergic neurons ; degeneration of these neurons represents an early pathological event in alzheimer 's disease . this protection mechanism through mglur7 activation is selectively inhibited by  -amyloid ( a ) .
a increases p21-activated kinase activity and decreases cofilin - mediated actin depolymerization through a p75(ntr)-dependent mechanism .
due to the lack of available ligands with specificity for mglur7 , mglur7-knockout mice were traditionally the only tool available to examine the physiological role of mglur7 and its involvement in cognitive , affective , and sensory behaviors .
mice lacking mglur7 receptors showed reduced short - term neural plasticity in the hippocampus   suggesting a role for mglur7 in the molecular mechanisms underlying cognition . in line with this evidence , mglur7-knockout mice displayed some impairments in learning and memory [ 58 - 60 ] .
specifically , these mice showed normal behavior in the t - maze   but impairments in both the 4-arm and 8-arm maze tasks , which require intact working memory capacity . intriguingly , after training , mglur7-knockout mice showed test performance similar to the their wild type counterparts , implying that training can overcome baseline differences in working memory capacity [ 58 , 59 , 61 ] . similarly , mglur7-deficient mice exhibited impairments in the morris water maze , although they achieved similar performance to wild type mice after training .
it has been thus proposed that mglur7 is involved in the molecular mechanisms underlying short - term working and spatial memory whereas long - term memory operates in an mglur7-independent manner [ 58 , 62 ] .
mice lacking mglur7 displayed delayed extinction of a conditioned fear response in the conditioned emotional responses ( cer ) procedure , but the initial acquisition of fear responses appeared unaltered in these mice [ 62 , 63 ] .
this may be due to the different neural substrates involved in acquisition ( the amygdala ) and extinction ( the hippocampus and prefrontal cortex ) of cer .
however , mglur7-deficient mice displayed reduced shock - induced freezing and impaired conditioned taste aversion , which are both amygdala - dependent paradigms [ 64 - 66 ] .
the reason for these discrepant findings may be the different experimental strategies used in the two studies .
the cer study used shocks that were signaled by auditory cues , and the conditioned taste aversion study used saccharin avoidance after pairing it with intraperitoneal injection of licl , an agent which causes transient visceral malaise .
mglur7-knockout mice exhibited reduced anxiety - like responses in several behavioral tests such as the elevated plus maze , staircase , marble burying , light - dark box , open field , and stress - induced hyperthermia tests .
in addition , and in accordance with their anxiolytic - like phenotype , mglur7 knockout mice also displayed reduced stress responses and activity of the hypothalamic - pituitary - adrenal ( hpa ) axis .
mglur7-deficient mice showed increased expression of hippocampal glucocorticoid and 5-hydroxytriptamine 1a receptors ( gr and 5-ht1a ) . increased gr and 5-ht1a expression in response to the lack of mglur7 suggests enhanced negative feedback of the hpa axis , which is hyperactive in depressant and anxiety-  like phenotypes [ 71 , 72 ] .
accordingly , mglur7-knockout mice showed increased hpa suppression in response to dexamethasone and increased levels of brain derived neurotrophic factor ( bdnf ) , both positive indexes of antidepressant activity [ 73 , 74 ] . consistently , mglur7-knockout mice exhibited antidepressant - like profiles in the tail suspension and forced swim tests [ 67 , 75 ] .
mice lacking the mglur7 receptor displayed an increased seizure vulnerability and reduced anticonvulsant effects of ( rs ) phosphonophenylglycine ( ppg ) , a broad spectrum group iii mglur agonist , indicating a potential neuroprotective role for mglur7 .
altogether , mglur7-knockout mouse strategies have largely contributed to uncovering the role of mglur7 in epilepsy , cognition , and emotion regulation .
one critical limitation of conventional knockout strategies , however , is that the constitutive lack of a gene may lead to compensatory effects from related proteins , especially during development , and this may confound some outcome measures . to address this concern ,
the development of selective mglur7 ligands for direct activation / blockade of the receptor in the adult organism is critical .
mglur7 displays low affinity for the  classic  group iii orthosteric agonists such as l - ap4 and l - sop .
moreover , orthosteric receptor activation requires an a - amino acid moiety and distal phosphonic group , which makes these compounds too hydrophilic to penetrate the blood - brain - barrier for subsequent brain exposure .
acpt-1 can penetrate the blood - brain - barrier , but has shown the same low selectivity for mglur7 [ 79 , 80 ] .
the competitive group iii mglur antagonist , ly341495 , shows the highest potency but is scarcely selective , since it is also a potent antagonist at group ii mglurs .
msop , cppg , and map4 are more selective for group iii mglurs but display weak potency [ 30 , 77 ] .
targeting allosteric binding sites has permitted drug developers to overcome the scarce selectivity and hydrophilicity associated with orthosteric compounds .
allosteric binding sites are less conserved among the other mglur family members and do not require hydrophilic moieties .
amn082 was developed as the first selective positive allosteric modulator ( pam ) for mglur7 .
amn082 fully activates mglur7 [ 78 , 84 ] , and its action is not blocked by mglur7 orthosteric antagonists .
amn082 , however , is rapidly metabolized into an active compound in vivo , which inhibits monoamine transporter activity .
consistent with the mglur7-knockout mouse phenotype , amn082 increases plasma corticosterone and adrenocorticotropic hormone ( acth ) levels .
amn082 has been shown to reduce fear acquisition and ltp in the amygdala , but improve fear extinction .
amn082 s effect on fear extinction is in line with the resistance to fear extinction observed in mglur7-deficient mice , whereas its effect on fear acquisition is divergent from the phenotype of mglur7-deficient mice , which displayed no abnormality in fear acquisition .
paradoxically produced anxiogenic - and anxiolytic - like effects [ 68 , 69 , 85 , 88 - 90 ] and also demonstrated antidepressant - like activity [ 89 , 91 , 92 ] , the latter being in contrast to the mglur7-knockout phenotype .
amn082 also facilitated nociception when microinjected into the ventrolateral pag ( vl pag ) , central nucleus of the amygdala ( cea ) , or nucleus tractus solitarius ( nts ) [ 40 , 88 , 93 ] .
amn082 changed the activity of neurons that respond to pain stimuli in the rostral ventromedial medulla ( rvm ) and decreased glutamate release into the vl pag , consistently with descending pathway inhibition and pain facilitation [ 40 , 94 - 96 ] .
amn082 has been shown to facilitate nociception in some studies ; however , it reduced pain responses in other studies [ 98 , 99 ] .
one possible explanation for the contradictory effects of amn082 is the rapid and long - lasting mglur7 receptor internalization induced by amn082 , which coincides with functional antagonism .
alternatively , amn082 s scarce selectivity  for mglur7 in vivo suggests the possibility of off - target involvement [ 83 , 100 ] .
amn082 also reduces ethanol and cocaine intake [ 101 , 102 ] , facilitates the extinction of aversive memories , and increases colonic secretory function .
the recent discovery of novel negative allosteric modulators ( nams ) for mglur7 will likely contribute to better understanding of the functional role  for mglur7 in neural functioning .
6-(4-methoxyphenyl)-  5-methyl-3-pyridin-4-ylisoxazolo[4,5-c]pyridin-4(5h)-one ( mmpip ) , a selective negative allosteric modulator for mglur7 , has shown inverse agonist activity for mglur7 and good brain exposure after systemic administration [ 103 , 104 ] . in vivo studies with mmpip
have shown that negative allosteric modulation of mglur7 impairs cognitive performance in the object recognition and radial arm maze tasks and reduces social interaction in rodents .
mmpip has been found to be ineffective in a range of behavioral experiments aimed at investigating motor coordination , anxiety and depression - like behaviors , sensorimotor gating , seizure threshold , and nociception in healthy rodents .
later , a recent paper from our laboratory confirmed that , when administered into the vl pag , mmpip showed no effect in healthy rats but inhibited pain responses in formalin and neuropathic pain models .
mmpip altered pain thresholds by modulating the antinociceptive descending pathway at rvm levels when administered into the vl pag .
specifically , in neuropathic rats it increased the activity of antinociceptive off cells and decreased that of pronociceptive on cells , consistently with antinociception , but proved ineffective in healthy controls .
this context - dependent mmpip effect was confirmed in a novel study where mmpip was shown to reverse the main symptoms of neuropathic pain in the spared nerve injury model , while remaining ineffective in control mice . in neuropathic mice
, systemic mmpip administration increased thermal and mechanical thresholds , occurrence of open - arm choice in the elevated plus maze , reduced immobility in the tail suspension test , and reduced the number of marbles buried and digging events in the marble - burying test , thus demonstrating putative anxiolytic- and antidepressant - like properties .
changes in receptor expression in some supraspinal areas such as the basolateral amygdala , dorsal raphe , prelimbic cortex , pag , and hippocampus have been observed in neuropathic conditions .
in line with the efficacy of mmpip in chronic pain models and its lack of effect in healthy animals , mmpip showed context - dependent activity in recombinant cell lines and inactivity under normal physiological conditions .
other selective mglur7 nams have recently been developed : 7-hydroxy-3-(4-iodophenoxy)-4h - chromen-4-one ( xap044 ) , which has shown to inhibit long - term potentiation ( ltp ) in the lateral amygdala in brain slices from wild type mice but not in mglur7 knockout mice , thus suggesting xap044 specific action on mglur7 . in vivo experiments
have shown that xap044 is brain penetrant and , similar to mglur7 knockout mice , produces anti - stress , antidepressant- , and anxiolytic - like effects and reduces freezing in a fear conditioning paradigm .
a single systemic xap044 administration reverted mechanical allodynia and ameliorated anxiety- and depression - like behaviors in a model of neuropathic pain .
interestingly , ( + ) -6-(2,4-dimethylphenyl)-2-ethyl-6,7-dihydrobenzo [ d]oxazol-4(5h)-one ( adx71743 ) , another selective mglur7 nam , has demonstrated excellent brain exposure after subcutaneous administration and anxiolytic - like effects in the elevated plus maze and marble burying tests .
the recent development of selective mglur7 nams has profoundly contributed to the delineation of a functional role for mglur7 in physiological and pathological conditions . apart from providing a better understanding of mglur7 function at the synapse
indeed mmpip , xap044 , and adx71743 have demonstrated selectivity for mglur7 and behavioral effects in line with the mglur7-knockout phenotype .
mmpip and xap044 have also been tested in chronic pain conditions and co - morbid affective and cognitive disorders , thus their possible therapeutic exploitation is reasonable .
a summary of mglur7 positive and negative allosteric modulators effects is presented in table 1 .
the greater importance of mglur7 of all mglurs is unveiled by its wide distribution and its high evolutionary conservation . in particular ,
mglur7 exhibits high expression in excitatory and inhibitory synapses within the brain , which are considered critical for neurologic and pathologic disorders .
initial studies using mglur7-knockout mice have suggested that mglur7 is involved in a series of neurological and psychiatric disorders such us epilepsy , anxiety , and depression .
these effects were later confirmed by similar findings obtained using selective mglur7 allosteric modulators . moreover , some of the novel mglur7 negative allosteric modulators , apart from clarifying the function of mglur7 in fine tuning excitatory and inhibitory synapses within the cns , have confirmed the mglur7-knockout phenotype and clarified the role of mglur7 in neuropsychiatric disorders . in this context , mmpip and xap044 have also been tested in models of neuropathic pain and have shown promising anti - allodynic , anti - anxiety- , and anti - depressant - like effects .
further , these compounds have been reported to improve cognitive performance , which is deeply affected in these models of chronic pain . as a direct consequence of these findings , further studies investigating mglur7
negative allosteric modulator effects are expected to facilitate development of novel therapeutics for pain and pain - related affective and cognitive disorders .
 metabotropic glutamate receptor 7 ( mglur7 ) is localized presynaptically at the active zone of neurotransmitter release . 
 unlike mglur4 and mglur8 , which share mglur7 s presynaptic location , mglur7 shows low affinity for glutamate and is activated only by high glutamate concentrations . 
 its wide distribution in the central nervous system ( cns ) and evolutionary conservation across species suggest that mglur7 plays a primary role in controlling excitatory synapse function . 
 high mglur7 expression has been observed in several brain regions that are critical for cns functioning and are involved in neurological and psychiatric disorder development . until the recent discovery of selective ligands for mglur7 , techniques to elucidate its role in neural function were limited to the use of knockout mice and gene silencing . 
 studies using these two techniques have revealed that mglur7 modulates emotionality , stress and fear responses . 
 n , n'-dibenzhydrylethane-1,2-diamine dihydrochloride ( amn082 ) was reported as the first selective mglur7 allosteric agonist . 
 pharmacological effects of amn082 have not completely confirmed the mglur7-knockout mouse phenotype ; this has been attributed to rapid receptor internalization after drug treatment and to the drug s apparent lack of in vivo selectivity . 
 therefore , the more recently developed mglur7 negative allosteric modulators ( nams ) are crucial for understanding mglur7 function and for exploiting its potential as a target for therapeutic interventions . 
 this review presents the main findings regarding mglur7 s effect on modulation of synaptic function and its role in normal cns function and in models of neurologic and psychiatric disorders . 