The role of NMDA receptor-operated calcium channels in persistent nociception after formalin-induced tissue injury.

The contribution of intracellular calcium to central sensitization and persistent nociception in response to tissue injury in rats was examined following the subcutaneous injection of formalin into the hindpaw. Formalin injury-induced nociceptive behaviors were enhanced by intrathecal pretreatment with the calcium ionophore A23187 or the calcium channel agonist Bay-K8644. Conversely, formalin nociceptive responses were reduced by intrathecal pretreatment with the calcium chelator Quin 2 or the calcium channel antagonists verapamil and nifedipine. Each of these agents affected the tonic, but not the acute, phase of the formalin response. The enhancement in formalin nociceptive behavior in rats treated with L-aspartate or L-glutamate was reversed by combined pretreatment with the noncompetitive NMDA antagonist MK-801, but not by nifedipine or the non-NMDA excitatory amino acid antagonist 6-cyano-7-dinitroquinoxaline-2,3-dione. In rats not treated with excitatory amino acids, the analgesic effect of MK-801 was also significantly greater than that produced by nifedipine. Furthermore, combining nifedipine with MK-801 did not produce a significantly greater analgesic effect than MK-801 alone. The results suggest that central sensitization and persistent nociception following formalin-induced tissue injury are dependent on the influx of calcium through predominantly NMDA receptor-operated (and to a lesser extent voltage-gated) calcium channels.     

Intracellular Messengers Contributing to Persistent Nociception and Hyperalgesia Induced by L-Glutamate and Substance P in the Rat Formalin Pain Model

The contribution of the intracellular messengers nitric oxide, arachidonic acid and protein kinase C to persistent nociception in response to tissue injury in rats was examined following the subcutaneous injection of formalin into the hindpaw. Formalin injury-induced nociceptive behaviours were reduced by intrathecal pretreatment with inhibitors of nitric oxide synthase (N^G-nitro-_L-arginine methyl ester, _L-NAME), arachidonic acid (dexamethasone) or protein kinase C [protein kinase C (19–26) and 1–95-(isoquinolinesulphonyl)-2-methylpiperazine dihydrochloride, H-7]. Each of these agents affected the tonic, but not the acute, phase of the formalin response. Furthermore, none of these agents affected mechanical or thermal flexion reflex thresholds in rats not injected with formalin. Conversely, formalin-induced nociceptive responses were enhanced by stimulators of nitric oxide (sodium nitroprusside), arachidonic acid metabolism (arachidonic acid) or protein kinase C [(±)-1-oleoyl-2-acetyl-glycerol], and were slightly reduced by inositol trisphosphate. Mechanical flexion reflexes were also reduced by arachidonic acid, while thermal flexion reflexes were reduced after treatment with sodium nitroprusside, arachidonic acid or [(±)-1 -oleoyl-2-acetyl-glycerol]. The enhancement of formalin nociceptive behaviours (hyperalgesia) in rats treated with _L-glutamate or substance P was reversed by pretreatment with inhibitors of nitric oxide (_L-NAME), arachidonic acid (dexamethasone) or protein kinase C (H-7). The results suggest that central sensitization and persistent nociception following formalin-induced tissue injury, and the hyperalgesia in the formalin test induced by _L-glutamate and substance P, are dependent on the intracellular messengers nitric oxide, arachidonic acid and protein kinase C.     

Behavioral sensitization to kainic acid and quisqualic acid in mice: comparison to NMDA and substance P responses.

Substance P (SP) and the excitatory amino acid (EAA) agonists NMDA, kainic acid (KA), or quisqualic acid (Quis) each produce a transient, caudally directed biting and scratching response (CBS) in mice after their intrathecal injection. We have previously shown that repeated injections of SP result in a decrease in the intensity of CBS, or desensitization. The goals of the present study were (1) to determine whether desensitization also develops to the CBS behavior produced by EAAs in the spinal cord, (2) to characterize the role of interneurons in desensitization, and (3) to examine possible interactions between EAAs and SP. While injection of NMDA at 2 min intervals resulted in desensitization to its CBS behavioral effect, behavioral responses to repeated injections of KA or Quis increased in intensity, exhibiting sensitization. The NMDA antagonist DL-2-amino-5-phosphonovaleric acid failed to alter sensitization to either KA or Quis but inhibited behaviors produced by SP and NMDA, suggesting an NMDA-mediated component in SP-induced behavior. Concanavalin A, which is reported to block desensitization to the electrophysiologic effect of Quis, blocked sensitization to the behavioral effects of both Quis and KA. Strychnine, bicuculline, and 5-aminovaleric acid each inhibited desensitization to SP and NMDA, supporting the notion of recruitment of inhibitory transmitters in the attenuation of NMDA and SP activity. Pretreatment with capsaicin selectively inhibited the development of behavioral sensitization to KA, suggesting an involvement of small-diameter C-fibers in the enhancement of responsivity to KA. Consistent with this, pretreatment with SP selectively potentiated the CBS response to KA. The potentiation of KA effects by SP and dependence of KA behavioral sensitization on C-fiber activity suggest a possible mechanism by which EAAs and SP may be involved in the mediation of pain.     

Potent Analgesia Induced in Rats by Combined Action at PCP and Polyamine Recognition Sites of the NMDA Receptor Complex

The present study was performed to examine the analgesic effects of the intrathecal administration of agents acting at various sites in the N -methyl- d -aspartic acid (NMDA) receptor complex on the nociceptive responses to s.c. formalin injection in rats. Both the competitive NMDA receptor antagonist 2-amino-5-phosphonovaleric acid (APV) and the non-competitive NMDA antagonist dizocilpine maleate (MK-801) produced dose-dependent analgesic effects in the late, but not the early, phase of the formalin test. The polyamine antagonist ifenprodil, and the strychnine-insensitive glycine antagonists DCQX and 7-chlorokynurenic acid, failed to produce any analgesic effects in either the early or the late phase of the formalin test. The analgesic effects of APV were enhanced slightly by combined administration with a non-analgesic dose of glycine, and the analgesic effects of MK-801 were dramatically potentiated by combined adminstration of a non-analgesic dose of the polyamine spermine. The results indicate that much more potent analgesia can be produced in the formalin test by a combination of open channel blockers (such as MK-801) with agonists acting at the polyamine site, than by a single treatment with antagonists to either glycine allosteric or polyamine sites within the NMDA receptor complex.     

Excitatory amino acids: role in morphine excitation in rat periaqueductal gray

Morphine was previously found to elicit an explosive excitatory behavior following its injection at a high dose in the rat periaqueductal gray (PAG). This non-naloxone reversible excitatory action of morphine was mimicked by the GABAA receptor antagonist, bicuculline, suggesting that morphine excitation was due in part to GABAA receptor blockade. In this paper, we report that injections of the excitatory amino acid (EAA) analogues, N-methyl-D-aspartate (NMDA), quisqualate (Q) or kainate (K) in the rat PAG resulted in similar (but not identical) behaviors. The excitatory actions of morphine or of NMDA (but not Q or K) were blocked or attenuated by the NMDA receptor antagonist, 2-amino-7-phosphonoheptanoate. These results show that both GABAA receptors as well as receptors for the EAAs may contribute to the excitatory actions of morphine in the PAG, and suggest that GABA may normally function to counterbalance a tonic excitatory influence of the EAAs.     

Unilateral subcutaneous bee venom but not formalin injection causes contralateral hypersensitized wind-up and after-discharge of the spinal withdrawal reflex in anesthetized spinal rats

This study aimed to investigate the effect of tonic nociception on spinal withdrawal reflexes including (1) long lasting spontaneous responses elicited by subcutaneous (s.c.) administration of formalin (2.5%, 50 microl) and bee venom (BV, 0.2 mg/50 microl) into the hind paw and (2) corresponding ipsilateral (primary) and contralateral (secondary) hypersensitivity to noxious pinch and repetitive supra-threshold (1.5 x T) electrical stimuli at different frequencies (3 Hz: wind-up; 20 Hz: after-discharge) in anesthetized spinal rats. Spinal withdrawal reflexes were studied by simultaneously assessing single motor units (SMUs) electromyographic (EMG) activities from the bilateral medial gastrocnemius (MG) muscles. Subcutaneous formalin-induced persistent spontaneous SMU EMG responses were in typical biphasic manner with an apparent silent period (about 13-18 min), but in contrast, BV elicited monophasic long lasting (about 1 h) SMU EMG responses without any resting state. The mechanically and electrically evoked responsiveness of SMUs were enhanced significantly by ipsilateral BV injection, whereas enhanced electrically, but not mechanically, evoked responses (including wind-up and after-discharge) were found at the non-injection site of the contralateral hind paw. However, s.c. administration of formalin was only able to establish ipsilateral hypersensitivity of the SMUs to repeated electrical, not mechanical, stimulation. Neither mechanically nor electrically evoked contralateral hypersensitivity of the SMUs was found during the ipsilateral formalin-induced nociception. For pharmacological intervention, intrathecal administration of the non-N-methyl-d-aspartate (non-NMDA) receptor antagonist CNQX (40 nmol/10 microl), but not the non-competitive NMDA receptor antagonist MK-801 (40 nmol/10 microl), significantly depressed BV-induced contralateral hypersensitivity of the SMUs to repeated 3 Hz (wind-up) and 20 Hz (after-discharge) frequencies of electrical stimulation. Using the extracellular SMU recording technique, we found that s.c. administration of formalin and BV shows a significant difference in long lasting spontaneous firing of SMUs. This is consistent with previous observations in animal behavioral studies. Additionally, contralateral electrically evoked hypersensitivity of the SMUs was found only following BV injection, not in the formalin test. The maintenance and development of BV-induced contralateral hypersensitivity of the spinal withdrawal reflex to noxious electrical stimulation indeed depend on different central pharmacological receptors. The spinal non-NMDA, but not the NMDA, receptors may play important role in BV-induced contralateral central hyperexcitability and sensitization.     

Differential effect of peripheral glutamate (NMDA,non-NMDA) receptor antagonists on bee venom-induced spontaneous nociception and sensitization

This study aimed to investigate the role of peripheral N-methyl-d-aspartate (NMDA) and non-NMDA receptor on (1). spontaneous nociception and (2). on sensitization induced by subcutaneous (s.c.) injection of bee venom (0.2mg/50 micro l) in rats. Peripheral s.c. administration of the competitive NMDA receptor antagonist dl-2-amino-5-phosphonovaleric acid (AP5), the non-competitive NMDA receptor channel blocker MK-801, and the competitive non-NMDA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) were performed before (pre-treatment) and after (post-treatment) bee venom-induced inflammation. Pre-treatment with AP5 (10mM, 50 micro l) and both pre-treatment and post-treatment with MK-801 (2mM, 50 micro l) into the same area of the bee venom injection site markedly reduced the bee venom-increased spontaneous responses of wide-dynamic range (WDR) neuron of the spinal cord. Post-treatment with the same dose of AP5 as well as pre-treatment and post-treatment with CNQX (5mM, 50 micro l) did not produce any inhibitory effects. Additionally, the role of peripheral NMDA and non-NMDA receptors on bee venom-induced mechanical allodynia and hyperalgesia were investigated and assessed by the paw withdrawal reflex to the innocuous and noxious mechanical stimulation. Peripheral administration of AP5, but not CNQX, reduced mechanical allodynia and hyperalgesia. The data suggest that the peripheral NMDA receptor, but not non-NMDA receptor, plays a pivotal role in the bee venom-induced persistent nociception and hyperexcitability.     

Glutamate receptor antagonists block cocaine-induced convulsions and death.

The involvement of excitatory amino acid (EAA) receptors in mediation of the toxic effects of cocaine was studied in male ICR mice. Cocaine HCl (90 mg/kg, IP) induced seizures in 95% and death within 24 h in 68% (n = 135) of the animals. There was a significant correlation (r = .54) between the time to onset of convulsions and the time to death in mice which died within 30 min of injection (n = 84). Pretreatment with selected EAA receptor antagonists 15 min prior to cocaine differentially blocked cocaine toxicity. Selective N-methyl-D-aspartic acid (NMDA) receptor antagonists (MK-801, dextrorphan, CPP) decreased both the incidence of seizures and mortality. A nonselective EAA antagonist, kynurenic acid, decreased lethality in doses which did not reduce convulsions. A similar action was observed following pretreatment with the selective kainic acid/AMPA receptor antagonist, GDEE. Antagonists at EAA receptors can provide significant protection against cocaine-induced toxicity. Moreover, the data provide evidence for the involvement of both NMDA and non-NMDA receptor subtypes in aspects of cocaine toxicity.     

Glutamate receptor regulation of rat nucleus accumbens neurons in vivo

Extracellular single cell recording and microiontophoretic techniques were used to characterize the roles of ionotropic and metabotropic glutamate receptors (iGluRs and mGluRs) in glutamate-induced excitation of rat nucleus accumbens (NAc) neurons in vivo. Pulse-ejected glutamate (16–128 nA) induced a current-dependent increase in the firing of quiescent NAc neurons. A stronger excitatory response to α-amino-3-hydroxy-5-methyl-4-iosoxazole-proprionic acid (AMPA) was observed at much lower ejection currents (0.1–6.4 nA). Compared to AMPA and glutamate, N-methyl-D -aspartate (NMDA) induced a much less potent excitation in a narrow current range (1–4 nA) and only when neurons were previously “primed” with other excitatory amino acids (EAAs). Higher ejection currents of all three EAA agonists drove NAc neurons into a state of apparent depolarization block. AMPA-evoked firing was selectively blocked by the AMPA receptor antagonist 6,7-dinitroquinoxaline-2,3-dione (DNQX) whereas NMDA-induced activity was selectively prevented by the NMDA receptor antagonist 2-amino-5-phosphonovalerate (D-AP5). DNQX, but not D-AP5, significantly attenuated glutamate-evoked activity. The mGluR receptor agonist (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid (1S,3R-t-ACPD) failed to evoke activity of NAc neurons, but significantly reduced the excitatory effects of other EAAs. This modulatory effect of 1S,3R-t-ACPD was consistently blocked by the selective mGluR antagonist L(+)-2-amino-3-phosphonopropionic acid (L-AP3) whereas another mGluR antagonist (RS)-4-carboxy-3-hydroxy phenylglycine (4C3HPG) was inconsistent in this regard. These results indicate that the excitatory effects of glutamate on rat NAc neurons in vivo are primarily mediated by non-NMDA iGluRs and that mGluRs function to dampen excessive glutamate transmission through iGluRs. © 1996 Wiley-Liss, Inc.     

Ca2+/calmodulin‐dependent protein kinase II in spinal dorsal horn contributes to the pain hypersensitivity induced by γ‐aminobutyric acid type a receptor inhibition

The fast inhibitory synaptic transmission mediated by the γ‐aminobutyric acid type A receptor (GABA_AR) within spinal dorsal horn exerts a gating control over the synaptic conveyance of nociceptive information from the periphery to higher brain regions. Although a large body of evidence has demonstrated that the impairment of GABAergic inhibition alone is sufficient to elicit pain hypersensitivity in intact animals, the underlying mechanisms remain to be characterized. The present study shows that Ca²⁺/calmodulin‐dependent protein kinase II (CaMKII) is an important signaling protein downstream of reduced GABAergic inhibition. We found that pharmacological removal of inhibition by intrathecal application of the GABA_AR antagonist bicuculline significantly enhanced the autophosphorylation of CaMKII at Thr286 in spinal dorsal horn of mice. In addition to increased CaMKII activity, bicuculline also promoted CaMKII interaction with N‐methyl‐D‐aspartate (NMDA)‐subtype glutamate receptors and induced the translocation of CaMKII from cytosolic compartments to the synaptosomal membrane fraction. Immunoblotting analysis revealed that the phosphorylation levels of NMDA receptor NR2B subunit at Ser1303 and of AMPA‐subtype glutamate receptor GluR1 subunit at Ser831, two important CaMKII phosphorylation sites, were substantially enhanced after bicuculline application. Behavioral tests illustrated that intrathecal administration of the CaMKII inhibitor KN‐93, NMDA receptor antagonist D‐APV, or AMPA receptor antagonist GYKI 52466 effectively ameliorated the mechanical allodynia evoked by bicuculline. These data thus indicate that CaMKII signaling is critical for the reduced inhibition to evoke spinal sensitization. © 2013 Wiley Periodicals, Inc.     

Quinolinate and kainate neurotoxicity in neostriatal cultures is potentiated by co-culturing with neocortical neurons

It has been suggested that a disorder in the regulation of excitatory amino acids (EAA) may underlie the loss of neostriatal neurons seen in Huntington's disease. The role of neocortical afferent fibers in determining the EAA sensitivity of neostriatal neurons was assessed by comparing EAA toxicity in co-cultures of neocortex and neostratum with that of neostriatum alone. In culturesalone, EAAs produced only modest neuronal losses. Kainate, which tended to be the most potent excitotoxin, produced a loss of approximately 30% of the neurons after a 5-min exposure at a 1-mM concentration. In co-cultures, the sensitivity of neostriatal neurons to EAA toxicity was dramatically enhanced; toxicity was increased about two-fold for kainate and quinolinate at millimolar concentrations and as much as 8-fold for quinolinate at micromolar concentrations. The effects of EAA co-incubation with the N-methyl-d-aspartate (NMDA) receptor antagonist, 2-amino-5-phosphonovaleric acid, suggested that the toxic actions of quinolinate, but not kainate, were mediated largely by NMDA receptors.     

Suppressing calcium/calmodulin-dependent protein kinase II activity in the ventral tegmental area enhances the acute behavioural response to cocaine but attenuates the initiation of cocaine-induced behavioural sensitization in rats

In the present experiments we administered an alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptor antagonist (CNQX), N-methyl-D-aspartate (NMDA) receptor antagonist (AP-5), or l-type calcium channel blocker (diltiazem) directly into the ventral tegmental area (VTA) before each of four daily systemic cocaine injections in order to assess their influence on the initiation phase of behavioural sensitization. Results indicated that pretreatment with CNQX or AP-5 impaired the initiation of cocaine-induced behavioural sensitization. Intra-VTA administration of diltiazem significantly increased the behavioural activation induced by an acute cocaine injection, but impaired the development of cocaine-induced behavioural sensitization. Because AMPA and NMDA receptors, as well as l-type calcium channels are calcium permeable, we also investigated the role of the calcium-activated second messenger calcium/calmodulin-dependent protein kinase II (CaM-KII). Similar to the results obtained with diltiazem, administration of the CaM-KII inhibitor KN-93 into the VTA enhanced the acute behavioural response to cocaine but prevented the augmentation of cocaine-induced behavioural hyperactivity following repeated injections. Consistent with this finding, the behavioural hyperactivity produced by cocaine was markedly enhanced among homozygous alpha-CaM-KII knockout mice but the initiation of behavioural sensitization to cocaine was attenuated relative to wild-type mice. Separate experiments performed in rats demonstrated an increase in total protein levels of CaM-KII in the VTA 24 h after the last of seven daily injections of cocaine. Taken together, these results indicate that blocking l-type calcium channels or impairing CaM-KII activity in the VTA augments the acute behavioural hyperactivity induced by cocaine. The present findings also suggest that increased calcium influx through AMPA receptors, NMDA receptors and l-type calcium channels on dopaminergic neurons in the VTA contributes significantly to the initiation of behavioural sensitization by amplifying calcium signalling through CaM-KII.     

NMDA receptor antagonists, MK-801 and ACEA-1011, prevent the development of tonic pain following subcutaneous formalin

Subcutaneous injection of formalin produces a biphasic pain response: an early, transient phase followed by a late tonic phase. The present study examined the involvement of the acid (NMDA) receptor in the development of the late pain produced following subcutaneous injection of formalin into the hind paw in mice. Blockade of the NMDA receptor by its non-competitive antagonist, MK-801, prior to formalin injection, but not after, reduced pain during the late phase. Similarly, blockade of the NMDA receptor allosteric site by the novel glycine site antagonist, ACEA-1011, also reduced the pain response in the late phase. These results suggest that the development of the late phase of formalin pain is due to NMDA-mediated activity during the early phase.     

Administration of either non-NMDA receptor agonists or NMDA receptor antagonists into the substantia nigra or the globus pallidus reduces the psychostimulant effect of high helium pressure on locomotor activity in rats

Helium pressure of >2 MPa is a well known factor underlying pressure-dependent central neuroexcitatory disorders that include locomotor and motor activity (LMA) and myoclonia. We investigated the effects of bilateral injection in either the substantia nigra (SN) or the globus pallidus (GP) of the AMPA receptor agonist (+/-)AMPA, the kainate receptor agonist kainic acid, the NMDA receptor agonist (+/-)-cis-piperidine-2,3-dicarboxylic acid (PDA), and the NMDA receptor antagonist (+/-)-2-amino-7-phosphono-heptanoic acid (AP-7) in the occurrence of helium pressure-induced LMA and myoclonia. Administration of AMPA, kainate, or AP-7 in either the SN or the GP significantly reduced high helium pressure-induced LMA, whereas the NMDA receptor agonist showed no significant effect. Injection in the SN of the non-NMDA receptor agonist AMPA and the NMDA receptor agonist PDA increased the development of high helium pressure-induced myoclonia, whereas injection of the NMDA receptor antagonist AP-7 into the SN or the GP decreased it. This confirms that NMDA transmission in the SN and the GP would play a major role in the development of helium pressure-induced LMA; manipulation of AMPA and kainate systems may have therapeutic potential. The opposite effects of AMPA on LMA and myoclonia also confirm the neural substrates involved in the motor disorder produced by helium pressure differ substantially between LMA and myoclonia.     

Effects of intracisternal injection of interleukin-6 on nociceptive jaw opening reflex and orofacial formalin test in freely moving rats

The present study was performed to investigate the effects of central cytokines on the modulation of nociception in the orofacial area. To achieve this purpose, a nociceptive jaw opening reflex and an orofacial formalin test were monitored before and after intracisternal administration of interleukin-6 (IL-6) in freely moving rats. In the nociceptive jaw opening reflex, the digastric electromyogram (dEMG) was not significantly changed after intracisternal injection of 200 pg and 2 ng IL-6. However, 20 ng IL-6 suppressed dEMG to 74+/-7% of the control values. In the inflammatory orofacial formalin test, intracisternal injection of 200 pg and 2 ng IL-6 did not change the number of noxious behavioral responses produced by formalin injection. However, 20 ng IL-6 injected intracisternally significantly increased the number of noxious behavioral responses produced by formalin. The hyperalgesic action of intracisternal IL-6 in the orofacial formalin test was blocked by pretreatment with interleukin-1 (IL-1) receptor antagonist. These results suggest that IL-6 injected intracisternally modulates the transmission of nociceptive information in the orofacial area. However, the hypo/hyper-algesic response of central cytokines seems to depend on the orofacial pain model. The hyperalgesic response of central IL-6 seems to be mediated by the IL-1 receptor.     

AIDA reduces glutamate release and attenuates mechanical allodynia after spinal cord injury

Spinal cord injury (SCI) leads to an increase in extracellular excitatory amino acid (EAA) concentrations, resulting in glutamate receptor-mediated excitotoxicity and central sensitization. To test contributions of group I metabotropic glutamate receptors (mGluRs) in SCI induced release of glutamate and in behavioral outcomes of central sensitization following injury, we administered 1-aminoindan-1,5-dicarboxylic acid (AIDA; 0.1 nmol intraspinally), a potent group I mGluR antagonist, to rats immediately after spinal cord contusion injury. EAAs were collected by microdialysis and quantified using HPLC. AIDA significantly decreased extracellular glutamate but not aspartate concentrations and significantly attenuated the development of mechanical but not thermal allodynia. These results suggest mGluRs play an important role in injury-induced EAA release and in central sensitization following SCI.     

NMDA receptors in nucleus accumbens modulate stress-induced dopamine release in nucleus accumbens and ventral tegmental area

Converging evidence suggests that dopamine (DA) transmission in nucleus accumbens (NAcc) is modulated locally by an excitatory amino acid (EAA)-containing input possibly originating in medial prefrontal cortex (PFC). In the present study, we examined the effects of intra-NAcc administration of EAA receptor antagonists on stress-induced increases of NAcc DA levels and of dendritically released DA in the ventral tegmental area (VTA). Local injection of the NMDA receptor antagonist—AP-5 (0.05, 0.5, and 5.0 nmoles)—dose-dependently potentiated increases in NAcc DA levels elicited by 15 min of restraint stress. In contrast, local application of equivalent doses of the kainate/AMPA receptor antagonist—DNQX—failed to alter the NAcc DA stress response reliably. In a separate experiment, we found that intra-NAcc injection of AP-5 also potentiated stress-induced increases in VTA DA levels. These results indicate that EAAs acting at NMDA receptors in NAcc can modulate stress-induced DA release in this region. Our data indicate, however, that this action exerts an inhibitory influence on the NAcc DA stress response, suggesting that the relevant population of NMDA receptors are not located on NAcc DA terminals. The fact that intra-NAcc AP-5 injections also potentiated the DA stress response in VTA suggests instead an action mediated by NMDA receptors located on NAcc neurons that feedback, directly or indirectly, to cell bodies of the mesocorticolimbic DA system. Synapse 26:225–234, 1997. © 1997 Wiley-Liss Inc.     

Blocking NMDA receptors in the hippocampal dentate gyrus with AP5 produces analgesia in the formalin pain test.

The hippocampus is an integral component of the "limbic" system and, as such, may contribute to the negative affect and avoidance motivation experienced during pain. A substantial body of evidence indicates that the hippocampus processes pain-related information, that some hippocampal neurons respond exclusively to painful stimulation, and that long-term anatomical changes occur in dentate gyrus neurons, following noxious physical stimulation. NMDA receptor antagonist drugs administered to the hippocampus interfere with long-term potentiation, learning, and memory; these same drugs, when applied to the spinal cord, prevent the long-term neurophysiological changes caused by noxious physical stimulation. This experiment tested whether blocking NMDA receptors in the hippocampal formation reduces nociceptive behaviors in an animal model of persistent human pain. The competitive NMDA receptor antagonist AP5 was injected into the dentate gyrus of alert, unrestrained rats either 5 min before or 15 min following the administration of a subcutaneous injection of formalin irritant. Pain behaviors in both acute and tonic phases of the formalin test were significantly reduced by AP5 treatments. These results support the hypothesis that the hippocampal formation is involved in pain-related neural processing and that NMDA receptor-sensitive mechanisms in the hippocampus are involved in pain perception and/or the expression of pain-related behaviors.     

Effects of Excitatory Amino Acid Receptor Agonists and Antagonists on the Secretion of Melatonin, Luteinizing Hormone and Prolactin in the Ram

To assess the role of excitatory amino acids (EAA) as neurotransmitters in the transmission of light information from the retina to the pineal gland, we have determined whether the systemic injection of EAA agonists in Soay rams will mimic the suppressive effect of light on the secretion of melatonin, and whether pretreatment of rams with EAA antagonists will block this effect. In addition, the efficacy of the drugs in affecting neuroendocrine systems was investigated by measuring the changes in the secretion of luteinizing hormone (LH) and prolactin. Injections of the EAA receptor agonist, NMDA (N-methyl-D,L-aspartate: 4.0 mg/kg iv), and the non-NMDA type EAA receptor agonist, AMPA (DL-α-amino-3-hydroxy-5-methylisoxazole-propionic acid : 0.2 mg/kg iv) given at night to rams exposed to long days (16 h light:8 h darkness), caused no change in the blood plasma concentrations of melatonin. The treatments induced an acute increase in the concentrations of LH, and NMDA, but not AMPA, caused a sustained increase in the concentrations of prolactin. Injections of the specific NMDA-type receptor antagonist, CGP (CGP 37849: 1.0 mg/kg iv) and the non-NMDA-type receptor antagonist, DNQX (6,7 Dinitroquinoxaline-2,3-dione: 0.5 mg/kg iv), given prior to a 1-h light period at night, in rams under long days, caused no change in the light-induced decrease in blood plasma concentrations of melatonin. The drug treatments had no effect on the plasma concentrations of LH, but CGP, and not DNQX, stimulated an acute increase in the plasma concentrations of prolactin. These results provide support for the hypothesis that EAA mechanisms operate in the hypothalamus to regulate the release of peptides and catecholamines which control the secretion of LH and prolactin from the pituitary gland; different sub-types of EAA receptors are involved in the control of the two pituitary hormones. The failure of the treatments to affect the secretion of melatonin may indicate that EAA receptor activation is not involved in the photic relay to the pineal gland, or may merely reflect the inability of the drugs to penetrate into the retina/SCN/pineal neural circuits to produce a response.     

NMDA receptors in the anterior cingulate cortex mediate pain-related aversion

Using a rat formalin-induced conditioned place avoidance (F-CPA) model and Fos immunohistochemistry, the present study observed the effect of N-methyl-D-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methyl-isoxozole propionic acid/kainite (AMPA/KA) receptors on pain-related aversion. Adult Sprague-Dawley rats were implanted with cannula in the anterior cingulate cortex (ACC) or the lateral ventricle. Before (10 min) the injection of formalin into a hindpaw on days 2 and 4 of place-conditioning trials, vehicle (0.01 M PBS), the NMDA receptors antagonist, 2-amino-5-phosphonovalerate (AP5), or the AMPA/KA receptors antagonist, 6,7-dinitroquinoxaline-2,3-dione (DNQX), was injected through the cannula. F-CPA was effectively eliminated by both intracerebroventricular (icv) and intra-ACC microinjection of AP5. In contrast, the intra-ACC or icv injection of DNQX failed to alter the conditioning scores of F-CPA compared with vehicle control group (P >0.05). Intra-ACC or icv injection of AP5 or DNQX had no effect on formalin-induced acute nociceptive behaviors. Fos immunoreactivity in the ACC was activated by retrieval of pain-related aversion, and this activation was significantly suppressed by preadministration of AP5, but not DNQX (P <0.001). These results suggest that NMDA receptors in the ACC are preferentially involved in the processing of the affective dimension of pain.