K. E. Binns

Contributions of mGlu1 and mGlu5 receptors to interactions with N-methyl-d-aspartate receptor-mediated responses and nociceptive sensory responses of rat thalamic neurons

The nociceptive responses of rat ventrobasal thalamus neurons can be reduced by N-methyl-D-aspartate antagonists and by selective metabotropic glutamate receptor mGlu1 antagonists. The recent development of the mGlu5-selective antagonist 6-methyl-2-(phenylethynyl)-pyridine now allows the direct probing of the possible involvement of mGlu5 receptors in thalamic nociceptive responses. Extracellular recordings were made from single neurons in the ventrobasal thalamus and immediately overlying dorsal thalamic nuclei of adult urethane-anaesthetized rats using multi-barrel electrodes. Responses of neurons to iontophoretic applications of the mGlu5-selective agonist (R,S)-2-chloro-5-hydroxyphenylglycine were selectively reduced during continuous iontophoretic applications of 6-methyl-2-(phenylethynyl)-pyridine. Similar applications of 6-methyl-2-(phenylethynyl)-pyridine reduced neuronal responses to noxious thermal stimuli to 53+/-9.5% of control responses. Co- application by iontophoresis of N-methyl-D-aspartate and metabotropic glutamate receptor agonists resulted in a mutual potentiation of excitatory responses. This effect could be reduced by either 6-methyl-2-(phenylethynyl)-pyridine or the mGlu1 antagonist LY367385. These results, taken together with previous data, suggest that acute thalamic nociceptive responses are mediated by a combination of mGlu1, mGlu5 and N-methyl-D-aspartate receptor activation, and that co-activation of these receptors produces a synergistic excitatory effect. Thus blockade of any of these receptor types would have a profound effect on the overall nociceptive response.

Different roles for GABAA and GABAB receptors in visual processing in the rat superior colliculus

1 The superficial grey layer of the superior colliculus (SGS) contains a high proportion of GABAergic inhibitory neurones. We have investigated the role of GABA receptors in synaptic transmission of aspects of visual activity in the SGS that may be driven by inhibitory mechanisms, such as surround inhibition and response habituation. 2 Multi‐barrel glass iontophoretic pipettes were used to record single neuronal activity in the SGS of urethane‐anaesthetized rats. Visual stimulation was provided by the display of moving bars and stationary spots of light on a monitor placed in the receptive field. 3 Both ejection of GABA and the GABAB agonist baclofen reduced responses to moving bars (interstimulus intervals ≥ 8 s). The effects of GABA were reversed by the GABAA antagonist bicuculline, and the effects of baclofen were antagonized by the GABAB antagonist CGP 35348. 4 Surround inhibition was estimated by plotting the response to flashed spots of increasing diameter. In controls, expanding the spot diameter beyond the excitatory receptive field caused a decrease in the response. This inhibitory surround was reversibly reduced by bicuculline, but CGP 35348 had no effect. 5 Response habituation is the progressive reduction in the visual response during repetitive stimulus presentation. In controls, the visual response was reduced to 44 ± 3% of its initial level when a stimulus (moving bar) was presented 5 times with an interstimulus interval of 0.5 s. During CGP 35348 ejection, response habituation was reversibly reduced. Bicuculline had no effect on response habituation. 6 The effects of bicuculline on surround inhibition in the superior colliculus are consistent with similar studies in the lateral geniculate nucleus which indicate that GABAA receptors mediate this effect. The function of GABAB receptors in the visual system is less well researched. The reduction of response habituation with CGP 35348 demonstrates that, at least in the SGS, GABAB receptors have an important role in visual transmission which is distinct from that of GABAA receptors.

Antagonism of the mGlu5 agonist 2‐chloro‐5‐hydroxyphenylglycine by the novel selective mGlu5 antagonist 6‐methyl‐2‐(phenylethynyl)‐pyridine (MPEP) in the thalamus

Our previous work has shown that Group I mGlu receptors participate in thalamic sensory processing in vivo. However, unequivocal demonstration of mGlu5 participation has not been possible due to the lack of specific ligands. We have therefore made a preliminary study of the in vivo actions of the agonist (R,S)‐2‐Chloro‐5‐hydroxyphenylglycine [CHPG] and the novel mGlu5 antagonist 6‐methyl‐2‐(phenylethynyl)‐pyridine [MPEP] in order to characterize their suitability for functional studies. Iontophoretically administered MPEP selectively antagonized excitatory responses of single rat thalamic neurones to CHPG compared to the broad‐spectrum mGlu agonist (1S,3R)‐1‐aminocyclopentane‐1,3‐dicarboxylate. In contrast, the established mGlu1 and mGlu5 antagonist (S)‐4‐carboxyphenylglycine reduced responses to both agonists. These findings are the first demonstration of an in vivo action of CHPG and its antagonism by a selective mGlu5 antagonist. Furthermore MPEP appears to be a good tool for functional studies of mGlu5.

The functional influence of nicotinic cholinergic receptors on the visual responses of neurones in the superficial superior colliculus

In the rat, the superficial gray layer (SGS) of the superior colliculus receives glutamatergic projections from the contralateral retina and from the visual cortex. A few fibers from the ipsilateral retina also directly innervate the SGS, but most of the ipsilateral visual input is provided by cholinergic afferents from the opposing parabigeminal nucleus (PBG). Thus, visual input carried by cholinergic afferents may have a functional influence on the responses of SGS neurones. When single neuronal extracellular recording and iontophoretic drug application were employed to examine this possibility, cholinergic agonists were found to depress responses to visual stimulation. Lobeline and 1-acetyl-4-methylpiperazine both depressed visually evoked activity and had a tendency to reduce the background firing rate of the neurones. Carbachol depressed the visual responses without any significant effect on the ongoing activity, while the muscarinic receptor selective agonist methacholine increased the background activity of the neurones and reduced their visual responses. Lobeline was chosen for further studies on the role of nicotinic receptors in SGS. Given that nicotinic receptors are associated with retinal terminals in SGS, and that the activation of presynaptic nicotinic receptors normally facilitates transmitter release (in this case glutamate release), the depressant effects of nicotinic agonists are intriguing. However, many retinal afferents contact inhibitory neurones in SGS; thus it is possible that the increase in glutamate release in turn facilitates the liberation of GABA which goes on to inhibit the visual responses. We therefore attempted to reverse the effects of lobeline with GABA receptor antagonists. The depressant effects of lobeline on the visual response could not be reversed by the GABA(A) antagonist bicuculline, but the GABA(B) antagonist CGP 35348 reduced the effects of lobeline. We hypothesize that cholinergic drive from the parabigeminal nucleus may activate presynaptic nicotinic receptors on retinal terminals, thereby facilitating the release of glutamate onto inhibitory neurones. Consequently GABA is released, activating GABA(B) receptors, and thus the ultimate effect of nicotinic receptor activation is to depress visual responses.

Kainate receptor (GluR5)-mediated disinhibition of responses in rat ventrobasal thalamus allows a novel sensory processing mechanism

Kainate receptors have been studied extensively in vitro, but how they might function physiologically remains unclear. We studied kainate receptor modulation of synaptic responses in the rat ventrobasal thalamus using the novel antagonist LY382884 and the agonist ATPA (selective for GluR5‐containing kainate receptors) as tools. No evidence could be found for a direct contribution of kainate receptors to responses of thalamic relay cells to lemniscal (sensory) input in thalamic slices studied with the aid of intracellular and field potential recordings, using selective AMPA and NMDA receptor antagonists and LY382884. However, the GluR5 agonist ATPA reduced the IPSPs originating from the thalamic reticular nucleus. Extracellular single‐neurone recordings in anaesthetised rats showed that excitatory responses evoked by physiological vibrissa afferent stimulation were reduced by LY382884 applied iontophoretically at the recording site. This action of the antagonist was occluded when GABA receptors were blocked, indicating that the reduction in excitatory sensory responses by LY382884 is due to an action on GABAergic inhibition arising from the thalamic reticular nucleus. Further experiments showed that these actions depended on whether inhibition was evoked during activation of the excitatory receptive field rather than when inhibition was evoked from a surround vibrissa. We suggest that GluR5 is located presynaptically on inhibitory GABAergic terminals of thalamic reticular nucleus neurones, and that it is normally activated by glutamate spillover from synapses between excitatory afferents and relay neurones during physiological stimulation. We propose that this GluR5‐activated disinhibition has an important novel role in extracting sensory information from background noise.

Actions of the systemically active metabotropic glutamate antagonist MPEP on sensory responses of thalamic neurones

It is known that metabotropic glutamate receptors of the subtypes mGlu1 and mGlu5 participate in nociceptive processing in the thalamus, an area of prime importance in supra-spinal sensory processing. Antagonists of these receptors thus have potential as centrally-acting analgesics. We have investigated whether intravenous administration of the novel mGlu5-receptor antagonist 6-methyl-2-(phenylethynyl)-pyridine (MPEP) is able to reduce nociceptive responses of thalamic neurones. Extracellular recordings were made from single thalamic neurones of adult male Wistar rats anaesthetised with urethane. MPEP (1 mg/kg) reduced neuronal responses to noxious thermal stimuli to a mean of 24+/-4% of control within 10 min, whereas saline injections had no significant effect. Partial recovery was seen within 30-45 min after injection. Responses of neurones to non-noxious stimuli were not significantly affected by MPEP administration. In addition, MPEP caused an increase in the power of the slow-wave component (<1 Hz) of the electroencephalogram (EEG), but had no significant effect on peak frequency of the EEG or on heart rate. These results confirm that nociceptive responses of thalamic neurones are mediated in part by mGlu5 receptors. Furthermore, the effectiveness of intravenous MPEP suggests that such antagonists may be useful as centrally-acting analgesics.

Novel mode of nitric oxide neurotransmission mediated via S-nitroso-cysteinyl-glycine.

S‐nitroso‐cysteinyl‐glycine, a novel nitric oxide‐adduct thiol compound, can be detected in the brain (2.3 ± 0.6 pmol/mg protein), and released following stimulation of sensory afferents to the rat ventrobasal thalamus in vivo (resting conditions 17 n m; stimulation: 186 nm). Iontophoretic application of CysNOGly (20–80 nA) onto thalamic neurons in vivo resulted in enhancements of excitatory responses to either NMDA or AMPA (182 ± 13.6% and 244 ± 27.8% of control values, n = 15). CysNOGly enhanced responses to stimulation of vibrissal afferents to 132 ± 2.2% (n = 7) of control values. In contrast, the dipeptide CysGly reduced responses of ventrobasal neurons to NMDA and AMPA (54 ± 8.4% and 55 ± 10.8% of control, n = 5). CysNOGly was also a potent activator of soluble guanylate cyclase in vitro. Moreover, we found that NMDA elevated CysNOGly levels in vitro and this stimulatory effect was reduced by inhibitors of the neuronal NO synthase and of the γ‐glutamyl transpeptidase, suggesting that production of NO and CysGly is a prelude to CysNOGly synthesis. These findings suggest that the nitrosothiol CysNOGly plays a role in synaptic transmission in the ventrobasal thalamus. We propose a novel synaptic buffering mechanism where S‐nitroso‐cysteinyl‐glycine serves to restrict the locus of action of nitric oxide and so increase its local availability for target delivery. This could lead to a change in neuronal responses favouring sensory transmission similar to that seen in wakefulness or arousal in order to locally enhance transmission of persistent sensory stimuli.

Visual experience alters the molecular profile of NMDA-receptor-mediated sensory transmission.

N‐methyl‐d‐aspartate (NMDA) receptors (NMDArs) may facilitate experience‐dependent changes in the visual system. Early sensory experience has an influence over the production of the molecular components from which NMDArs are assembled, and thereby alters the properties of functional receptors. Using the antagonists d‐2‐amino‐5‐phosphonovalerate (AP5) and 3‐(2‐carboxypiperazin‐4‐yl)‐propyl‐1‐phosphonate (CPP), which have some selectivity for different variants of the NMDAr, we demonstrate that visual deprivation (by dark rearing) has functional consequences for NMDArs in the superior colliculus. An increase in the sensitivity of visual responses to AP5 in dark‐reared rats indicated that NMDArs were more important for visual transmission in these individuals. We also observed a relative change in the efficacy of the antagonists against the visual responses of normal versus dark‐reared rats. AP5 reduced the visual responses of both groups, but CPP was ineffective against visual responses after dark rearing. In the same neurons, CPP blocked NMDA induced activity indicating that molecular adaptations of NMDArs are specific to those synapses mediating visual activity.

Experience-dependent changes in the importance of N-methyl-d-aspartate (NMDA) receptors for visual transmission in superior colliculus

The excitatory amino acid transmitter glutamate mediates visual activity in the superficial grey layer (SGS) of superior colliculus. At eye opening N-methyl-d-aspartate receptors (NMDA-rs) convey little of the visual response, but with age their role in visual transmission increases to a peak at P21, then falls to the lower adult level. Visual deprivation which begins before eye opening causes NMDA-rs to assume a greater importance for visual transmission in SGS. Here we explore the possibility that these experience-dependent changes in the role of NMDA-rs in the SGS are limited by age. We find that the effects of visual deprivation on NMDA-r mediated visual activity are recoverable even after extensive dark rearing. Also, a short episode of visual experience is sufficient to allow the normal situation to be established and subsequent dark rearing is ineffective. Four-day periods of visual experience beginning at P14 or P25 have the same effect. Given that NMDA-rs take little part in visual transmission prior to P18, these data prompt a reconsideration of the role of NMDA-r mediated sensory transmission in the mechanisms by which early environmental experience influences the development of the visual system.