The Journal of Neurophysiology Vol. 80 No. 4 October 1998, pp. 2210-2214
Copyright ©1998 by the American Physiological Society

RAPID COMMUNICATION

₂-Adrenoceptors Modulate NMDA-Evoked Responses of Neurons in Superficial and Deeper Dorsal Horn of the Medulla

Kai-Ming Zhang, Xiao-Min Wang, Angela M. Peterson, Wen-Yan Chen, and Sukhbir S. Mokha

Department of Anatomy and Physiology, Meharry Medical College, Nashville, Tennessee 37208

	ABSTRACT

Abstract Introduction Methods Results Discussion References

Kai-Ming Zhang, Xiao-Min Wang, Angela M. Peterson, Wen-Yan Chen, and Sukhbir S. Mokha. ₂-Adrenoceptors modulate NMDA-evoked responses of neurons in the superficial and deeper dorsal horn of the medulla. J. Neurophysiol. 80: 2210-2214, 1998. Extracellular single unit recordings were made from neurons in the superficial and deeper dorsal horn of the medulla (trigeminal nucleus caudalis) in 21 male rats anesthetized with urethan. NMDA produced an antagonist-reversible excitation of 46 nociceptive as well as nonnociceptive neurons. Microiontophoretic application of a preferential ₂-adrenoceptor (₂AR) agonist, (2-[2,6-dichloroaniline]-2-imidazoline) hydrochloride (clonidine), reduced the NMDA-evoked responses of 86% (6/7) of nociceptive-specific (NS) neurons, 82% (9/11) of wide dynamic range (WDR) neurons, and 67% (4/6) of low-threshold (LT) neurons in the superficial dorsal horn. In the deeper dorsal horn, clonidine inhibited the NMDA-evoked responses of 94% (16/17) of NS and WDR neurons and 60% (3/5) of LT neurons. Clonidine facilitated the NMDA-evoked responses in 14% (1/17) of NS, 9% (1/11) of WDR, and 33% (2/6) of LT neurons in the superficial dorsal horn. Idazoxan, an ₂AR antagonist, reversed the inhibitory effect of clonidine in 90% (9/10) of neurons, whereas prazosin, an ₁-adrenoceptor antagonist with affinity for _2BAR, and _2CAR, were ineffective. We suggest that activation of ₂ARs produces a predominantly inhibitory modulation of the NMDA-evoked responses of nociceptive neurons in the medullary dorsal horn.

	INTRODUCTION

Abstract Introduction Methods Results Discussion References

The dorsal horn of the medulla (trigeminal nucleus caudalis) is an important relay for nociceptive and thermosensory information from the orofacial region (reviewed in Light 1992; Sessle 1987). Glutamate, a putative excitatory neurotransmitter (Watkins and Evans 1981), is present in trigeminal primary afferent fibers (Clements and Beitz 1991). Glutamate produces its actions by acting on modulate N-methyl-D-aspartic acid (NMDA), non-NMDA ionotropic [(±) -  - amino - 3 - hydroxy - 5 - methyl - isoxazole - 4 - propionic acid (AMPA)/Kainate], and metabotropic receptors in the spinal cord. The superficial dorsal horn of the medulla contains high densities of NMDA (Tallaksen-Greene et al. 1992) and non-NMDA ionotropic (Kondo et al. 1995; Tallaksen-Greene et al. 1992) and metabotropic (Tallaksen-Greene et al. 1992) receptors. NMDA receptors are involved in mediating nociceptive neurotransmission and neuroplasticity (hyperalgesia) in the dorsal horn (reviewed in Wilcox 1993). (2-[2,6-Dichloroaniline]-2-imidazoline) hydrochloride (clonidine), an ₂-adrenoceptor (₂AR) agonist, is reported to be effective in treating neuropathic pain in humans (Eisenach et al.1995; Tamsen and Gordh 1984) and in attenuating hyperalgesia induced by nerve injury or inflammation in animal studies (Kayser et al. 1995; Post et al. 1987; Puke et al.1994; Solomon et al. 1989; Xu et al. 1992). Autoradiographic studies demonstrated the presence of ₂ARs in the dorsal horn, particularly in the superficial dorsal horn of the spinal cord (Roudet et al. 1994; Unnerstall et al. 1984; Sullivan et al. 1987) and the medulla (Unnerstall et al. 1984). Microiontophoretically applied norepinephrine (NE) is reported to selectively inhibit nociceptive responses of deep dorsal horn neurons (Belcher et al. 1978; Fleetwood-Walker et al. 1985; Headley et al. 1978; reviewed in Jones 1991). Consistent with data from behavioral studies (reviewed in Jones 1991), the selective inhibitory effect of NE was suggested to involve ₂ARs (Davies and Quinlan 1985; Fleetwood-Walker et al. 1985). However, NE was also reported to produce nonselective inhibitory modulation of primate spinothalamic tract neurons (Willcockson et al. 1984) and interneurons in the superficial dorsal horn of the spinal cord in the rat (Howe and Zieglgänsberger 1987; Todd and Millar 1983). Although behavioral studies in mice indicated the importance of adrenoceptors in modulating biting and scratching behavior induced by intrathecal NMDA (Aanonsen and Wilcox 1987), no previous electrophysiological studies investigated the role of ₂-noradrenergic receptors in modulating the NMDA-evoked responses of physiologically characterized nociceptive neurons in the brain. This study was therefore designed to investigate the ₂AR-mediated modulation of NMDA-evoked responses of neurons in the superficial and deeper dorsal horn of the medulla.

	METHODS

Abstract Introduction Methods Results Discussion References

This study was based on data obtained from 21 male Sprague-Dawley rats (230-360 g) anesthetized with urethan (1.5 g/kg ip). Methods for animal preparation, monitoring the level of anesthesia, neuronal recording, and characterization of trigeminal neurons were similar to those described previously (Mokha 1992, 1993; Zhang et al. 1996). The rat's face was carefully shaved for adequate stimulation and mapping of receptive fields. To improve the stability of extracellular recordings from the superficial dorsal horn of the medulla, a small metallic brass plate was fixed to the skull, and the head was ventroflexed. The dorsal surface of the medulla was exposed and covered with warm agar (3-4% agar in normal saline at 40°C). All protocols were approved by the Institutional Animal Care and Use Committee of Meharry Medical College.

Recordings were made with the central barrel (2 M NaCl or 2% pontamine sky blue in 0.5 M sodium acetate) of a seven-barrel microelectrode (tip diameter, 4-10 µm; impedance, 2-15 M, Medical Systems). One barrel of the seven-barreled glass microelectrode contained 2 M NaCl for automatic current balancing, and the remaining barrels were filled with drug solutions for microiontophoretic application with the Neurophore BH2 system (Medical Systems). The solutions were either made up fresh before an experiment or thawed from a frozen state. The drug barrels contained solutions of the following drugs: NMDA (50 mM in 150 mM NaCl, pH 8.2), DL-2-amino-5-phosphonovaleric acid (AP-5, 50 mM in 150 mM NaCl, pH 8.0), clonidine (100 mM in double-distilled water, pH 4.5), idazoxan hydrochloride (100 mM in double-distilled water, pH 4.5), and (1-[4-amino-6,7-dimethoxy-2-quinazolinyl]-4-[2-furanyl-carbonyl]piperazine) hydrochloride (prazosin, 1.5 mM in double-distilled water, pH 4.5). All chemicals were obtained from Sigma Chemical. All drugs were ejected with positive current except NMDA and AP-5, which were ejected with negative current. Retaining currents were adjusted between 2 and 10 nA to prevent drug diffusion. Application of microiontophoretic currents 140 nA through a vehicle (150 mM NaCl, pH 4.5 and/or pH 8.0) containing barrel did not alter significantly the responses of neurons.

Single unit extracellular recordings were made from neurons in the superficial (laminae I and II) and deeper dorsal horn of the medulla. The procedures for recording from neurons in the superficial versus the deeper dorsal horn and the responses and receptive fields of neurons were described and discussed previously (Mokha 1992, 1993; Zhang et al. 1996). In addition, recording sites were also marked by electrophoretic ejection of pontamine sky blue (10 µA for 3 min) from the central recording barrel or one of the outer barrels as illustrated in Fig. 2B. Action potentials were recorded and amplified with conventional means and were continuously monitored on an oscilloscope to allow for discrimination of single units. Application of NMDA at low current was used to search for single units. Brush, pressure, pinch, squeeze, and radiant heat stimuli were used to characterize the responses of neurons to receptive field stimulation. Neurons were classified as selectively nocireceptive [nociceptive specific (NS), responding only to noxious stimuli], multireceptive [wide dynamic range (WDR), responding to both noxious and innocuous stimuli], and nonnociceptive [low threshold (LT), responding only to innocuous stimuli] as described previously (Mokha 1992, 1993; Zhang et al. 1996). Neural activity was fed to a window discriminator and to a microcomputer to allow analysis of the data on-line and off-line (MI² software). The histogram data files were transferred into ASCII files for further analysis with Microsoft Excel, SigmaStat, and SigmaPlot for Windows. The number of spikes was counted before, during, and after drug ejection. Stable control responses evoked by cyclical administration of NMDA (5-10 s on, 5-50 s off) were compared with responses during drug application. The effect of a test drug was defined as inhibitory or excitatory only when the NMDA-evoked responses differed from the mean of the control response by ±2 SD in the same direction. The time to one-half recovery (t_1/2) after the termination of the drug application was calculated. Grouped data are presented as the mean ± SE. Statistical analysis was performed by using paired t-test and one-way analysis of variance followed by Student-Newman-Keuls test. A probability level of P < 0.05 was considered significant.

View larger version (26K): [in this window] [in a new window]   FIG. 2. A: summary of the peak inhibitory effect of clonidine on the NMDA-evoked responses of neurons. : average control responses evoked by NMDA. : NMDA-evoked responses during the peak inhibitory effect of clonidine. y-axis (spikes/epoch = spikes/NMDA application) represents the total number of spikes evoked by NMDA application. Data presented as mean ± SE. Numbers below the bar represent number of neurons tested. Stars represent statistical significance (*P < 0.05; **P < 0.01; ***P < 0.001, paired t-test). B: location of recording sites in the superficial (laminae I and II) and deeper dorsal of the medulla. sp5: spinal trigeminal tract; Sp5C: spinal trigeminal nu, caudal; I and II: laminae I and II; pyx: pyramidal decussation.

	RESULTS

Abstract Introduction Methods Results Discussion References

Forty-six neurons were recorded extracellularly from the superficial (7 NS, 11 WDR, and 6 LT) and deeper (1 NS, 16 WDR, and 5 LT) dorsal horn. All neurons had an ipsilateral receptive field, and spontaneous activity was essentially absent. Responses evoked by NMDA were related to the intensity of the microiontophoretic current and displayed a short latency (<5 s). NMDA-evoked firing outlasted the ejection period by 3-10 s. AP-5 (10-30 nA), a selective NMDA receptor antagonist, blocked the NMDA-evoked responses in 100% of neurons (6/6) tested.

Clonidine (30 ± 3 nA, n = 41) reduced the NMDA-evoked responses of 86% (6/7) of NS neurons, 82% (9/11) of WDR neurons, and 67% (4/6) of LT neurons in the superficial dorsal horn (Figs. 1 and 2). It produced a peak inhibitory effect of 80 ± 4% (n = 19, P < 0.0001). Clonidine facilitated (153 ± 98%, n = 4, P < 0.05) the NMDA-evoked responses of 14% (1/7) of NS neurons, 9% (1/11) of WDR neurons, and 33% (2/6) of LT neurons in the superficial dorsal horn. In the deeper dorsal horn, clonidine reduced the NMDA-evoked responses of 94% (16/17) of NS and WDR neurons and 60% (3/5) of LT neurons. It produced a peak inhibitory effect of 71 ± 5% (n = 19, P < 0.0001). Biphasic effects were observed in 9% (1/11) of WDR neurons in the superficial dorsal horn and 40% (2/5) of LT neurons in the deeper dorsal horn. The inhibitory effect of clonidine exhibited longer lasting (t_1/2 = 2,262 ± 285 s, n = 5, range 1,710-3,345 s) or shorter lasting (t_1/2 = 255 ± 48 s, n = 19, range 35-800 s) time course. There was no significant correlation (P > 0.05, n = 24) between the time course and the dosage of clonidine (iontophoretic current × duration of application). Idazoxan, administered at currents that had no effect (P > 0.05, n = 10) when given alone, reduced the magnitude and/or the time course of the inhibitory effect of clonidine in 9 of 10 neurons (Fig. 1C). Idazoxan produced a significant reduction of 62 ± 10% in the time course of the inhibitory effect of clonidine (P < 0.05, n = 10, paired t-test). Significant reduction in the inhibitory effect of clonidine by idazoxan was also revealed in the combined data on the time course and the peak inhibitory effect of clonidine (P < 0.05, n = 10, paired t-test). Idazoxan reduced the peak inhibitory effect of clonidine by 69 ± 15% in 6 of 10 neurons. Prazosin, administered at currents that had no effect when given alone, did not alter the effect of clonidine in two of two neurons tested (Fig. 1B).

View larger version (29K): [in this window] [in a new window]   FIG. 1. 2-adrenoceptor activation mediates the inhibitory effect of clonidine on the N-methyl-D-aspartic acid (NMDA)-evoked responses of a wide dynamic range (WDR, excited by brush and pinch applied to the cutaneous receptive field) neuron in the superficial dorsal horn of the medulla. A: (2-[2,6-Dichloroaniline]-2-imidazoline) hydrochloride (clonidine) reduced the NMDA-evoked responses. B and C: inhibitory effect of clonidine was antagonized by idazoxan and not by prazosin. Neither prazosin nor idazoxan administered alone affected the NMDA-evoked responses.

	DISCUSSION

Abstract Introduction Methods Results Discussion References

This is the first in vivo electrophysiological demonstration of the ₂AR-mediated modulation of the NMDA-evoked responses of physiologically characterized neurons in the brain. The results presented here demonstrate that iontophoretically applied clonidine, an ₂AR agonist, produces a predominantly inhibitory modulation of the NMDA-evoked responses of nociceptive and nonnociceptive trigeminal neurons. This is consistent with the previously reported observations demonstrating the predominantly inhibitory modulation of glutamate-evoked responses by microiontophoretically applied NE in the dorsal horn of the spinal cord (Howe and Zieglgänsberger 1987; Willcockson et al. 1984). Inhibitory modulation of the NMDA-evoked responses of nociceptive neurons may contribute to the antinociceptive actions of clonidine observed in human (Eisenach et al. 1995; Tamsen and Gordh 1984) and animal studies (Kayser et al. 1995; Post et al. 1987; Solomon et al. 1989; Xu et al. 1992). NMDA receptor activation is intimately involved in mediating nociceptive neurotransmission and neural plasticity (hyperalgesia) after nerve injury or inflammation (reviewed in Wilcox 1993). Clonidine also facilitated the NMDA-evoked responses of some neurons in the superficial dorsal horn. This is consistent with previously reported observations on the glutamate-evoked responses of neurons in the superficial dorsal horn of the spinal cord (Millar and Williams 1989; Millar et al. 1993). Neurons facilitated by clonidine may represent a population of small inhibitory interneurons that can gate the transmission of nociceptive signals. The population of neurons facilitated by clonidine in our investigation is much smaller than that reported previously (Millar and Williams 1989; Millar et al. 1993). Several factors, such as the characteristics of recording electrodes and the iontophoretic paradigms, may contributed to this discrepancy. Although LT neurons are encountered less frequently in the superficial dorsal horn, nevertheless these are reported to be present in the superficial dorsal horn. Some of the superficial LT neurons in our study may include recordings made from neurons in the deeper dorsal horn because histological localization was not obtained for all LT neurons.

The modulation of NMDA-evoked responses by clonidine was significantly reduced by idazoxan, ₂AR antagonist. The ₂ARs exist in three pharmacologically distinguishable receptor subtypes (_2A, _2B, and _2C) (Bylund et al. 1994). Prazosin, which has high affinity for the ₁, _2B, and _2C adrenoceptors, did not alter the effects of clonidine on the NMDA-evoked responses. It is therefore likely that the _2A subtype of ₂ARs is involved in modulating the NMDA-evoked responses of neurons in the medullary dorsal horn. This is consistent with the previous anatomic findings indicating that the _2AAR appears to be the dominant receptor subtype present in neurons in the spinal dorsal horn (Nicholas et al. 1993; reviewed in Nicholas et al. 1996; Uhln and Wikberg 1991). In the medullary dorsal horn, _2AARs have been shown to be present predominantly in neurons in the superficial dorsal horn (Nicholas et al. 1993; Talley et al. 1996), whereas the _2C ARs are primarily located on neurons in the descending tract of the trigeminal nerve (Rosin et al. 1996). Receptor subtype dominance appears to be species specific because in the human spinal cord the _2B appears to be the dominant adrenoceptor (Smith et al. 1995). Evidence obtained recently in mouse strains with either a point mutation in the _2AAR gene or null mutation in the _2B and _2CAR genes demonstrated clearly that agonists at the ₂ARs produce antinociceptive action by acting primarily at the _2AARs (Hunter et al. 1997; Lakhlani et al. 1997; Stone et al. 1997).

Agonists at the ₂ARs which are imidazoline (clonidine, idazoxan) or imidazole (dexmedetomidine) also display moderate to high affinity binding to imidazole or imidazoline binding sites (reviewed in Codd et al. 1995). However, the antinociceptive action of clonidine in the spinal cord (Monroe et al. 1995) or PAG stimulation (Peng et al. 1996) was reported to be mediated by its action at the ₂ARs rather than on imidazoline receptors. Lack of analgesic effect of ₂ agonists in mice with a point mutation in the _2AAR gene strongly argues against the role of imidazoline receptors in contributing to the analgesic effects of ₂ agonists (Hunter et al. 1997; Lakhlani et al. 1997; Stone et al. 1997).

₂ARs are G-protein (G_i/G_o) coupled receptors and produce their actions by reducing the calcium conductance and enhancing the potassium conductance. Catecholamine terminals have been shown to make postsynaptic contacts with spinothalamic tract neurons (Westlund et al. 1990). Activations of ₂ARs increases potassium conductance in dorsal horn neurons, which produces hyperpolarization, decreases excitability, and contributes to analgesia (Grudt et al. 1995; North and Yoshimura 1984; Ocana and Baeyens 1993). Similar postsynaptic hyperpolarization induced by increased potassium conductance might contribute to the inhibitory modulation of NMDA-evoked responses observed in this investigation. This suggestion is consistent with previous observations reporting a nonselective inhibitory action of NE and clonidine in the dorsal horn (Höwe and Zieglgänsberger 1987; Peng et al. 1996; Willcockson et al. 1984). NE acting at ₁ARs was reported to produce -aminobutyric acid (GABA)-mediated inhibitory postsynaptic potentials in dorsal horn neurons (Grudt et al. 1995); nevertheless it is possible that clonidine-induced facilitations might be produced by postsynaptic inhibition of neurons containing GABA or glycine. Although direct axo-axonic contacts of noradrenergic terminals were not demonstrated on primary afferent fiber terminals (Doyle and Maxwell 1991a,b), electrophysiological studies suggest the involvement of presynaptic mechanisms in mediating the actions of NE and ₂ agonists (Jeftinija et al. 1981; Travagli and Williams 1996). Further, mRNA for ₂ARs (_2A, _2B, and _2C) is present in dorsal root ganglion cells (Gold et al. 1997; reviewed in Nicholas et al. 1996), and ganglionectomies reduce the binding of ₂ARs in the spinal cord. Activation of ₂ARs in dorsal root ganglion cells inhibits the N-type calcium channels (Cox and Dunlap 1992), which will be expected to reduce neurotransmitter release. Indeed, ₂ agonists are known to inhibit the stimuli-evoked release of substance P (Kuraishi et al. 1985; Pang and Vasko 1986) and glutamate (Kamisaki et al. 1993; Ueda et al. 1995) in the spinal cord. Therefore antinociceptive actions of ₂ agonists presumably involve both presynaptic and postsynaptic mechanisms.

The long-lasting (t_1/2 = 2,262 ± 285 s, n = 5, range 1,710-3,345) effects of clonidine observed in this investigation are consistent with previous observations in the spinal cord and the trigeminal system (Cahusac et al. 1995; Millar et al. 1993; Zhao and Duggan 1988). These longer-lasting effects presumably involve second messenger systems. Although the ₂ARs are known to reduce the production of cAMP in the spinal cord, raising levels of cAMP with forskolin did not alter the antinociceptive action of ₂ agonists (Uhln et al. 1990).

Predominantly inhibitory modulation of NMDA-evoked responses in the medullary dorsal horn may contribute to the antinociceptive effects of clonidine observed in human and animal studies.

	ACKNOWLEDGEMENTS

  We thank Dr. Jack Clark for critically reading the manuscript and allowing access to his image analysis facility, Dr. John S. Thomas for helping us with the statistical analysis of data, and D. Brown for reading an earlier draft of the manuscript.

  This work was supported by National Institutes of Health Grants DE-10903, RR-03032, DE-RR 10595, and MH-9843 and National Science Foundation Grants IBN-9109247 and HRD9255157.

	FOOTNOTES

  Address for reprint requests: S. S. Mokha, Dept. of Anatomy and Physiology, Meharry Medical College, 1005 D. B. Todd Blvd., Nashville, TN 37208.

  Received 23 March 1998; accepted in final form 3 July 1998.

	REFERENCES

Abstract Introduction Methods Results Discussion References

• AANONSEN, L. M., WILCOX, G. L. Nociceptive actions of excitatory amino acids in the mouse: effects of spinally administered opioids, phencyclidine and sigma agonists. J. Pharmacol. Exp. Ther. 243: 9-19, 1987.[Abstract/Free Full Text]
• BELCHER, G., RYALL, R. W., SCHAFFNER, R. The differential effects of 5-hydroxytryptamine, noradrenaline, and raphe stimulation on nociceptive and non-nociceptive dorsal horn interneurons. Brain Res. 151: 307-321, 1978.[Medline]
• BYLUND, D., EIKENBERG, D. C., HIEBLE, J., LLANZER, S. Z., LEFKOWITZ, R. J., MENNEMAN, K. P., MOLINOFF, P. B., RUFFOLO, R. R., TRENDELENBURG, U. International union of pharmacology nomenclature of adrenoceptors. Pharmacol. Rev. 46: 121-135, 1994.[Medline]
• CAHUSAC, P.M.B., MORRIS, R., HILL, R. G. A pharmacological study of the modulation of neuronal and behavioral nociceptive responses in the rat trigeminal region. Brain Res. 700: 70-82, 1995.[Medline]
• CLEMENTS, J. R., BEITZ, A. J. An electron microscopic description of glutamate-like immunoreactive axon terminal in the rat principal sensory and spinal trigeminal nuclei, J. Comp. Neurol. 309: 271-280, 1991.[Medline]
• CODD, E. E., PRESS, J. B., RAFFA, R. B. Alpha₂-adrenoceptors vs. imidazoline receptors: implications for ₂-mediated analgesia and other non-cardiovascular therapeutic uses. Life Sci. 56: 63-74, 1995.[Medline]
• COX, D. H., DUNLAP, K. Pharmacological discrimination of N-type from L-type calcium current and its selective modulation by transmitters. J. Neurosci. 12: 906-914, 1992.[Abstract]
• DAVIES, J., QUINLAN, E. Selective inhibition of responses of feline dorsal horn neurons to noxious cutaneous stimuli by tizanidine (DS103-282) and noradrenaline: involvement of ₂-adrenoceptors. Neuroscience 16: 673-682, 1985.[Medline]
• DOYLE, C. A., MAXWELL, D. J. Catecholaminergic innervation of the spinal dorsal horn: a correlated light and electron microscopic analysis of tyrosine hydroxylase-immunoreactive fibers in the cat. Neuroscience 45: 161-176, 1991a.[Medline]
• DOYLE, C. A., MAXWELL, D. J. Ultrastructural analysis of noradrenergic nerve terminals in the cat lumbosacral spinal dorsal horn: a dopamine--hydroxylase immunocytochemical study. Brain Res. 563: 329-333, 1991b.[Medline]
• EISENACH, J. C., DUPEN, S., DUBOIS, M., MIGUEL, R., ALLIN, D. Epidural clonidine analgesia for intractable cancer pain. Pain 61: 391-399, 1995.[Medline]
• FLEETWOOD-WALKER, S. M., MITCHELL, R., HOPE, P. J., MOLONY, V., IGGO, A. An ₂ receptor mediates the selective inhibition by noradrenaline of nociceptive responses of identified dorsal horn neurons. Brain Res. 334: 243-254, 1985.[Medline]
• GOLD, M. S., DASTMALCHI, S., LEVINE, J. D. Alpha (2)-adrenergic receptor subtypes in rat spinal dorsal root and superior cervical ganglion neurones. Pain 69: 179-190, 1997.[Medline]
• GRUDT, T. J., WILLIAMS, J. T., TRAVAGLI, R. A. Inhibition by 5-hydroxytryptamine and noradrenaline in substantia gelatinosa of guinea-pig spinal trigeminal nucleus. J. Physiol. (Lond.) 485: 113-120, 1995.[Abstract/Free Full Text]
• HEADLEY, P. M., DUGGAN, A. W., GRIERSMITH, B. T. Selective reduction by noradrenaline and 5-hydroxytryptamine of nociceptive responses of cat dorsal horn neurons. Brain Res. 145: 185-189, 1978.[Medline]
• HOWE, J. R., ZIEGLGÄNSBERGER, W. Responses of rat dorsal horn neurons to natural stimulation and to iontophoretically applied noradrenaline. J. Comp. Neurol. 255: 1-17, 1987.[Medline]
• HUNTER, J. C., FONTANA, D. J., HEDLEY, L. R., JASPER, J. R., LEWIS, R., LINK, R. E., SECCHI, R., SUTTON, J., EGLEN, R. M. Assessment of the role of ₂-adrenoceptor subtypes in the antinociceptive, sedative and hypothermic action of dexmedetomidine in transgenic mice. Br. J. Pharmacol. 122: 1339-1344, 1997.[Medline]
• JEFTINIJA, S., SEMBA, K., RANDIC, M. Norepinephrine reduces excitability of single primary afferent C-fibers in the cat spinal cord. Brain Res. 219: 456-463, 1981.[Medline]
• JONES, S. L. Descending noradrenergic influences on pain. In: Progress in Brain Research, edited by C. D. Barnes, and O. Pompeiano Amsterdam: Elsevier, 1991, vol. 88, p. 381-394.
• KAMISAKI, Y., HAMADA, T., MAEDA, K., ISHIMURA, M., ITOH, T. Presynaptic 2 adrenoceptors inhibit glutamate release from rat spinal cord synaptosomes. J. Neurochem. 60: 522-526, 1993.[Medline]
• KAYSER, V., DESMEULES, J., GUILBAUD, G. Systemic clonidine differentially modulates the abnormal reactions to mechanical and thermal stimuli in rats with peripheral mononeuropathy. Pain 60: 275-285, 1995.[Medline]
• KONDO, E., KIYAMA, H., YAMANO, M., SHIDA, T., UEDA, Y., TOHYAMA, M. Expression of glutamate (AMPA type) and -aminobutyric acid (GABA)_A receptors in the rat caudal trigeminal nucleus. Neurosci. Lett. 186: 169-172, 1995.[Medline]
• KURAISHI, Y., HIROTA, N., SATO, Y. Noradrenergic inhibition of the release of substance P from the primary afferents in the rabbit spinal dorsal horn. Brain Res. 359: 177-182, 1985.[Medline]
• LAKHLANI, P. P., MACMILLAN, L. B., GUO, T. Z., MCCOOL, B. A., LOVINGER, D. M., MAZE, M., LIMBIRD, L. E. Substitution of a mutant _2a-adrenergic receptor via "hit and run" gene targeting reveals the role of this subtype in sedative, analgesic, and anesthetic-sparing responses in vivo. Proc. Natl. Acad. Sci. USA 94: 9950-9955, 1997.[Abstract/Free Full Text]
• LIGHT, A. R. In: The Initial Processing of Pain and its Descending Control: Spinal and Trigeminal Systems. Basel, Switzerland: Karger, 1992.
• MILLAR, J., O'BRIEN, F. E., WILLIAMS, G. V., WOOD, J. The effects of iontophoretic clonidine on neurones in the rat superficial dorsal horn. Pain 53: 137-145, 1993.[Medline]
• MILLAR, J., WILLIAMS, G. V. Effects of iontophoresis of noradrenaline and stimulation of the periaqueductal gray on single-unit activity in the rat superficial dorsal horn. J. Comp. Neurol. 287: 119-133, 1989.[Medline]
• MOKHA, S. S. Differential influence of naloxone on the responses of nociceptive neurons in the superficial versus the deeper dorsal horn of the medulla in the rat. Pain 49: 405-413, 1992.[Medline]
• MOKHA, S. S. Morphine differentially modulates nociceptive input in the superficial versus the deeper dorsal horn of the medulla (trigeminal nucleus caudalis) in the rat. Brain Res. 626: 318-321, 1993.[Medline]
• MONROE, P. J., SMITH, D. L., KIRK, H. R., SMITH, D. J. Spinal noradrenergic imidazoline receptors do not mediate the antinociceptive action of intrathecal clonidine in the rat. J. Pharmacol. Exp. Ther. 273: 1057-1062, 1995.[Abstract/Free Full Text]
• NICHOLAS, A. P., HÖKFELT, T., PIERIBONE, V. A. The distribution and significance of CNS adrenoceptors examined with in situ hybridization. Trends Pharmacol. Sci. 17: 245-255, 1996.[Medline]
• NICHOLAS, A. P., PIERIBONE, V., HÖKFELT, T. Distribution of mRNA for alpha-2 adrenergic receptor subtypes in rat brain: an in situ hybridization study. J. Comp. Neurol. 328: 575-594, 1993.[Medline]
• NORTH, R. A., YOSHIMURA, M. The action of noradrenaline on neurons of the rat substantia gelatinosa in vitro. J. Physiol. (Lond.) 349: 43-56, 1984.[Abstract/Free Full Text]
• OCANA, M., BAEYENS, J. M. Differential effects of K⁺ channel blockers on antinociception induced by alpha-2-adrenoceptor, GABA_B and kappa-opioid receptor agonists. Br. J. Pharmacol. 110: 1049-1054, 1993.[Medline]
• PANG, I. H., VASKO, V. R. Morphine and norepinephrine but not 5-hydroxytryptamine and -aminobutyric acid inhibit the potassium stimulated release of substance P from rat spinal cord slices. Brain Res. 376: 268-278, 1986.[Medline]
• PENG, Y. B. lin, Q., AND WILLIS, W. D. Involvement of alpha-2 adrenoceptors in the periaqueductal gray-induced inhibition of dorsal horn cell activity in rats. J. Pharmacol. Exp. Ther. 278: 125-135, 1996.[Abstract/Free Full Text]
• POST, C., GORTH, T., MINOR, B., ARCHER, T., FREEDMAN, J. Antinociceptive and spinal cord tissue concentrations after intrathecal injection of guanfacine or clonidine into rats. Anesth. Analg. 66: 496-501, 1987.
• PUKE, M.J.C., LUO, L., XU, X.-J. The spinal analgesic role of ₂-adrenoceptor subtypes in rats after peripheral nerve section. Eur. J. Pharmacol. 260: 227-232, 1994.[Medline]
• ROSIN, D. L., TALLEY, E. M., LEE, A., STORNETTA, R. L., GAYLINN, B. D., GUYENET, P. G., LYNCH, K. R. Distribution of alpha 2C-adrenergic receptor-like immunoreactivity in the rat central nervous system. J. Comp. Neurol. 372: 135-165, 1996.[Medline]
• ROUDET, C., MOUCHET, P., FEUERSTEIN, C., SAVASTA, M. Normal distribution of alpha 2-adrenoceptors in the rat spinal cord and its modification after noradrenergic denervation: a quantitative autoradiographic study. J. Neurosci. Res. 39: 319-329, 1994.[Medline]
• SESSLE, B. J. The neurobiology of facial and dental pain: present knowledge, future directions. J. Dent. Res. 66: 962-981, 1987.[Abstract/Free Full Text]
• SMITH, M. S., SCHAMBRA, U. B., WILSON, K. H., PAGE, S. O., HULETTE, C., LIGHT, A. R., SCHWINN, D. A. ₂-Adrenergic receptors in human spinal cord: specific localized expression of mRNA encoding ₂-adrenergic receptor subtypes at four distinct levels. Mol. Brain Res. 34: 109-117, 1995.[Medline]
• SOLOMON, R. E., BRODY, M. J., GEBHART, G. F. Pharmacological characterization of alpha adrenoceptors involved in the antinociceptive and cardiovascular effects of intrathecally administered clonidine. J. Pharmacol. Exp. Ther. 251: 27-38, 1989.[Abstract/Free Full Text]
• STONE, L. S., MACMILLAN, L. B., KITTO, K. F., LIMBIRD, L. E., WILCOX, G. L. The _2a adrenergic receptor subtype mediates spinal analgesia evoked by ₂ agonists and is necessary for spinal adrenergic-opioid synergy. J. Neurosci. 17: 7157-7165, 1997.[Abstract/Free Full Text]
• SULLIVAN, A. F., DASHWOOD, M. R., DICKENSON, A. H. ₂-adrenoceptor modulation of nociception in rat spinal cord: location, effects and interactions with morphine. Eur. J. Pharmacol. 138: 169-177, 1987.[Medline]
• TALLAKSEN-GREENE, S. J., YOUNG, A. B., PENNEY, J. B., BEITZ, A. J. Excitatory amino acid binding sites in the trigeminal principal sensory and spinal trigeminal nuclei of the rat. Neurosci. Lett. 141: 79-83, 1992.[Medline]
• TALLEY, E. M., ROSIN, D. L., LEE, A., GUYENET, P. G., LYNCH, K. R. Distribution of alpha 2A-adrenergic receptor-like immunoreactivity in the rat central nervous system. J. Comp. Neurol. 372: 111-134, 1996.[Medline]
• TAMSEN, A., GORDH, T. E. Epidural clonidine produces analgesia. Lancet ii: 231-232, 1984.
• TODD, A. J., MILLAR, J. Receptive fields and responses to iontophoretically applied noradrenaline and 5-hydroxytryptamine of units recorded in laminae I-III of cat dorsal horn. Brain Res. 288: 159-167, 1983.[Medline]
• TRAVAGALI, R. A., WILLIAMS, J. T. Endogenous monoamines inhibit glutamate transmission in the spinal trigeminal nucleus of the guinea-pig. J. Physiology 491: 177-185, 1996.[Abstract/Free Full Text]
• UEDA, M., OYAMA, T., KURAISHI, Y., AKAIKE, A., SATOH, M. Alpha2-adrenoceptor-mediated inhibition of capsaicin-evoked release of glutamate from rat spinal dorsal horn slices. Neurosci. Lett. 188: 137-139, 1995.[Medline]
• UHLN, S., PERSSON, M.-L., ALARI, L., POST, C., AXELSSON, K. L., WIKBERG, J.E.S. Antinociceptive actions of ₂-adrenoceptor agonists in the rat spinal cord: evidence for antinociceptive ₂-adrenoceptor subtypes and dissociation of antinociceptive ₂-adrenoceptors from cyclic AMP. J. Neurochem. 55: 1905-1914, 1990.[Medline]
• UHLN, S., WIKBERG, J.E.S. Rat spinal cord ₂-adrenoceptors are of _2A-subtype: comparison with _2A- and _2B-adrenoceptors in rat spleen, cerebral cortex and kidney using [³H]RX821002 ligand binding. Pharmacol. Toxicol. 69: 341-350, 1991.[Medline]
• UNNERSTALL, J. R., KOPAJTIC, T. A., KUHAR, M. J. Distribution of ₂ agonist binding sites in the rat and human central nervous system: analysis of some functional, anatomic correlates of the pharmacologic effects of clonidine and related adrenergic agents. Brain Res. Rev. 7: 69-101, 1984.
• WATKINS, J. C., EVANS, R. H. Excitatory amino acid transmitters. Annu. Rev. Pharmacol. Toxicol. 21: 165-204, 1981.[Medline]
• WESTLUND, K. N., CARLTON, S. M., ZHANG, D., WILLIS, W. D. Direct catecholaminergic innervation of primate spinothalamic tract neurons. J. Comp. Neurol. 299: 178-186, 1990.[Medline]
• WILCOX, G. L. Spinal mediators of nociceptive neurotransmission and hyperalgesia. APS J. 2: 265-275, 1993.
• WILLCOCKSON, W. S., CHUNG, J. M., HORI, Y., LEE, K. H., WILLIS, W. D. Effects of iontophoretically released amino acids and amines on primate spinothalamic tract cells. J. Neurosci. 4: 732-740, 1984.[Abstract]
• XU, X.-J., PUKE, M.J.C., WIESENFELD-HALLIN, S. The depressive effect of intrathecal clonidine on the spinal flexor reflex is enhanced after sciatic nerve section in rats. Pain. 51: 145-151, 1992.[Medline]
• ZHANG, K.-M., WANG, X.-M., MOKHA, S. S. Opioids modulate N-methyl-D-aspartic acid (NMDA)-evoked responses of neurons in the superficial and deeper dorsal horn of the medulla (trigeminal nucleus caudalis). Brain Res. 719: 229-233, 1996.[Medline]
• ZHAO, Z.-Q., DUGGAN, A. W. Idazoxan blocks the action of noradrenaline but not spinal inhibition from electrical stimulation of the locus coeruleus and nucleus Kolliker-Fuse of the cat. Neuroscience 25: 997-1005, 1988.[Medline]

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