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The substantia gelatinosa (SG) of the trigeminal subnucleus caudalis (Vc; medullary dorsal horn) receives and processes orofacial nociceptive inputs, and serotonergic fibers involved in the descending modulation of nociception are more densely distributed in the superficial laminae of the Vc. This study investigated the direct effects of 5-HT(1A/7) receptor agonist 8-OH-DPAT on SG neurons of the Vc to assess functional expression of the 5-HT⁷ receptor using gramicidin-perforated patch-clamp in postnatal day (PND) 5-84 male mice. Of the 70 SG neurons tested, bath application of 8-OH-DPAT (30 μM) induced depolarization (n=33), hyperpolarization (n=16) or no response (n=21). In another 10 SG neurons, 8-OH-DPAT in the presence of 5-HT(1A) receptor antagonist WAY-100635 (1 μM) elicited either depolarization (n=6) or no response (n=4); hyperpolarization was not observed. The 8-OH-DPAT-induced depolarization was significantly blocked by the selective 5-HT⁷ receptor antagonist SB-269970 (10 μM; n=8), but not by WAY-100635 (1 μM; n=5). The depolarizing effect of 8-OH-DPAT was maintained in the presence of TTX, CNQX, AP5, picrotoxin, and strychnine, indicating direct postsynaptic action of 8-OH-DPAT on SG neurons (n=6). 5-HT⁷ receptor mRNA was also detected in five of 21 SG neurons by single-cell RT-PCR. The mean amplitude of 8-OH-DPAT-induced depolarization in PND 5-21 mice (n=21) was significantly larger than that in PND 22-84 mice (n=12), although the proportion of SG neurons responding to 8-OH-DPAT by depolarization did not differ significantly between two age groups of mice. These results indicate that 5-HT⁷ receptors are functionally expressed in a subpopulation of SG neurons of the Vc and activation of 5-HT⁷ receptors plays an important role in modulating orofacial nociceptive processing in the SG neurons of the Vc.

Serotonin (5-hydroxytryptamine, 5-HT) is involved in the descending modulation of nociceptive transmission in the spinal dorsal horn. The trigeminal subnucleus caudalis (Vc; medullary dorsal horn) processes nociceptive input from the orofacial region, and 5-HT-containing axons are numerous in the superficial layers of the Vc. This study examined the actions of 5-HT on the substantia gelatinosa (SG) neurons of the Vc, using gramicidin-perforated patch-clamp recording in brainstem slice preparations from immature mice. In order to clarify the possible mechanisms underlying 5-HT actions in the SG of the Vc, the direct membrane effects of 5-HT and effects of 5-HT receptor subtype agonists were examined. 5-HT induced a hyperpolarization in the majority (64/115, 56%) of the SG neurons tested. Thirty nine (34%) SG neurons showed no response, and 12 (10%) neurons responded with depolarization. The hyperpolarizing response to 5-HT was concentration-dependent (0.1-30 μM; n=7), not desensitized by repeated application (n=22), and significantly attenuated by Ba(2+) (K(+) channel blocker; n=8). The 5-HT-induced hyperpolarization was maintained in the presence of TTX (Na(+) channel blocker), CNQX (non-NMDA glutamate receptor antagonist), AP5 (NMDA glutamate receptor antagonist), picrotoxin (GABA(A) receptor antagonist), and strychnine (glycine receptor antagonist), indicating direct postsynaptic action of 5-HT on SG neurons (n=7). The 5-HT-induced hyperpolarizing effects were mimicked by 8-OH-DPAT (5-HT(1A) receptor agonist) and α-methyl-5-HT (5-HT(2) receptor agonist) and blocked by WAY-100635 (5-HT(1A) receptor antagonist) and ketanserin (5-HT(2) receptor antagonist). Single-cell RT-PCR also revealed the presence of mRNA for 5-HT(1A) and 5-HT(2C) subtypes in the SG neurons. These results suggest that 5-HT acts directly on SG neurons and 5-HT-induced hyperpolarization is mediated, in part, by 5-HT(1A) receptors and 5-HT(2) receptors, as well as by the activation of K(+) channels, indicating an important role for 5-HT in the modulation of orofacial nociceptive processing at the level of the SG of the Vc in mice.

The substantia gelatinosa (SG) of the trigeminal subnucleus caudalis (Vc) has been implicated in the processing of nociceptive information from the orofacial region. Kainate receptors (KARs) play an important role in sensory transmission. Five different KAR subunits have been cloned and the expression of the KAR subunits showed developmental changes. In this study, RT-PCR, western blotting, immunohistochemistry and a patch clamp technique were used examine the functional expression of the GluR5 subunit in the SG of the Vc in juvenile, peripubertal and/or adult mice. The levels of mRNA and protein expression of the GluR5 subunit in the SG of the Vc were higher in the juvenile mice than in the peripubertal or adult mice. In addition, the KA and ATPA, a GluR5 KAR agonist, induced membrane depolarization on the SG neurons in both juvenile and adult mice in a concentration-dependent manner. However, the juvenile SG neurons showed a stronger response to KA and ATPA than those of adults. The membrane depolarization by KA was suppressed slightly in the presence of the AMPA receptor antagonist, GYKI 52466. These results show that the GluR5 KAR subunits are expressed functionally on the SG neurons of the Vc in mice, and the expression levels of the GluR5 subunits decrease with postnatal development. These postnatal changes in the GluR5 KAR subunit may be a possible mechanism for age-dependent pain processing.

The substantia gelatinosa (SG) of the trigeminal subnucleus caudalis (Vc) receives many thin-myelinated Adelta-fiber and unmyelinated C primary afferent fibers and has been implicated in the processing of nociceptive information. Somatostatin (SST) is a neuromodulator in the brain and spinal cord. A number of studies have demonstrated that SST can play a key role in pain modulation at the spinal cord level. However, there is little information available on functional SST receptor expression in the SG neurons of the Vc in mice. This study examined the direct membrane effects of SST and SST receptor type 2 agonist, seglitide (SEG) on the SG neurons of Vc in gramicidin perforated current clamp mode. In addition, SSTR2 mRNA expression was detected on the SG neurons using single cell RT-PCR in juvenile mice. Most SG neurons (37/68, 54%) were hyperpolarized after a bath application of SST. When SST was applied in stages, the second responses (83% of the first response) were less intense than those after the first application suggesting that SSTRs are desensitized by repeated application. The SST-induced hyperpolarizing response was maintained in the presence of TTX (Na(+) channel blocker), AP-5 (NMDA receptor antagonist), CNQX (non-NMDA glutamate receptor antagonist), picrotoxin (GABA(A) receptor antagonist) and strychnine (glycine receptor antagonist), respectively, suggesting that SST has direct effects on the postsynaptic SG neurons. SSTR2 mRNA was detected in 11 out of 28 (39%) SG neurons tested. The SST-induced hyperpolarizing effects were mimicked by SEG, a SSTR2 agonist. These results suggest that functional SSTR2 receptors are expressed on the SG neurons of Vc in juvenile mice and can be a potential target for modulating orofacial pain.

The substantia gelatinosa (SG) of the trigeminal subnucleus caudalis (Vc; medullary dorsal horn) receives and processes orofacial nociceptive inputs, and serotonergic fibers involved in the descending modulation of nociception are more densely distributed in the superficial laminae of the Vc. This study investigated the direct effects of 5-HT(1A/7) receptor agonist 8-OH-DPAT on SG neurons of the Vc to assess functional expression of the 5-HT⁷ receptor using gramicidin-perforated patch-clamp in postnatal day (PND) 5-84 male mice. Of the 70 SG neurons tested, bath application of 8-OH-DPAT (30 μM) induced depolarization (n=33), hyperpolarization (n=16) or no response (n=21). In another 10 SG neurons, 8-OH-DPAT in the presence of 5-HT(1A) receptor antagonist WAY-100635 (1 μM) elicited either depolarization (n=6) or no response (n=4); hyperpolarization was not observed. The 8-OH-DPAT-induced depolarization was significantly blocked by the selective 5-HT⁷ receptor antagonist SB-269970 (10 μM; n=8), but not by WAY-100635 (1 μM; n=5). The depolarizing effect of 8-OH-DPAT was maintained in the presence of TTX, CNQX, AP5, picrotoxin, and strychnine, indicating direct postsynaptic action of 8-OH-DPAT on SG neurons (n=6). 5-HT⁷ receptor mRNA was also detected in five of 21 SG neurons by single-cell RT-PCR. The mean amplitude of 8-OH-DPAT-induced depolarization in PND 5-21 mice (n=21) was significantly larger than that in PND 22-84 mice (n=12), although the proportion of SG neurons responding to 8-OH-DPAT by depolarization did not differ significantly between two age groups of mice. These results indicate that 5-HT⁷ receptors are functionally expressed in a subpopulation of SG neurons of the Vc and activation of 5-HT⁷ receptors plays an important role in modulating orofacial nociceptive processing in the SG neurons of the Vc.