Effects of c-fos antisense oligodeoxynucleotide (ASO) on serotonin (5-HT)-induced upregulation of preprodynorphin (ppDyn), preproenkephalin (ppEnk), and glutamic acid decarboxylase (GAD), a special chemical marker for c-aminobutyric acid (GABA) neurons, mRNAs in cultured spinal dorsal horn neurons were investigated in order to extend our understanding of expressions of opioid peptides and GABA in spinal cord regulated by the descending serotonergic efferents. Reverse transcription-polymerase chain reaction analysis revealed a time-course increase in the expression of mRNAs encoding c-fos, ppDyn, ppEnk, and GAD after administration of 5-HT (100 nM). Administration of c-fos ASO (0.02 nM) 30 min prior to 5-HT application markedly blocked the expression of c-fos gene. Moreover, c-fos ASO pretreatment significantly decreased the 5-HT-induced upregulation of ppDyn and ppEnk mRNAs, but failed to affect the expression level of GAD mRNA. These results suggest that the serotoningic raphe-spinal efferents might play an important role in regulating the synthesis of enkephalin, dynorphin, and GABA in the spinal dorsal horn neurons. The immediate early oncogene c-fos might be involved in the 5-HT-induced increase in ppDyn and ppEnk expression. However, under the present experimental conditions, c-fos does not seem to be associated with the upregulation of GAD mRNA induced by 5-HT.

Injection of bee venom into one hindpaw of rat can elicit acute inflammation together with spontaneous pain, heat hyperalgesia and mechanical hyperalgesia/allodynia in the injected paw. 5-Hydroxytryptamine (5-HT)1A receptor is the predominant receptor subtype in the spinal dorsal horn mediating the function of 5-HT in nociception. The goal of the present study is to assess the role of 5-HT1A receptor in the pain associated with the bee venom induced inflammation. Here we showed that 1 or 4 h after a subcutaneous bee venom challenge, expression of 5-HT1A receptor mRNA in the ipsilateral lumbar spinal cord increased significantly by 80.94 or 37.86%, respectively. Antisense oligodeoxynucleotide knockdown of spinal 5-HT1A receptor attenuated spontaneous pain and reversed heat hyperalgesia in rats injected with bee venom. Thus, the present data suggest a facilitating role for 5-HT1A receptor in bee venom induced inflammatory pain.

The synaptic connections between g-aminobutyric acid (GABA)- and glycine-immunoreactive terminals and neurons projecting to the lateral parabrachial region were examined by a combination of retrograde tracing and immunohistochemical staining in the rat medullary dorsal horn. After injection of horseradish peroxidase (HRP) into the right lateral parabrachial region, HRP retrogradely labeled neurons were observed bilaterally in laminae I, II and III of the medullary dorsal horn with an ipsilateral predominance. GABA- and glycine-like immunoreactive terminals were found in laminae I, II and III. Some of these GABA- and glycine-like immunoreactive terminals were observed chiefly to make symmetric synapses with HRP-labeled neuronal cell bodies and dendritic processes. The present results indicate that neurons in the medullary dorsal horn projecting to the lateral parabrachial region might be modulated by GABAergic and glycinergic inhibitory intrinsic neurons, which might be significantly involved in the regulation of the noxious information transmission.

Morphology and electrical membrane properties of neurons in the superficial part of the magnocellular layer of the rat medullary dorsal horn (MDH: caudal subnucleus of the spinal trigeminal nucleus) were examined by using horizontal slice preparations. Intracellular recording and biocytin-injection combined with histochemical and immunohistochemical staining were done. Twenty-four neurons were examined successfully and classified into projection neurons (PNs) and intrinsic neurons (INs). The PNs were further divided into type I PNs (I-PNs) and type II PNs (II-PNs). The I-PNs sent axons to the medullary reticular formation; the II-PNs sent axons to the interpolar subnucleus of the spinal trigeminal nucleus but had no axons extending to the medullary reticular formation. The INs that sent no axons to the brain regions outside the MDH were also divided into small INs with spiny dendrites (INSSs) and large INs with aspiny dendrites (INLAs). The dendritic fields of the PNs extended to laminae I and II of the MDH and occasionally further to the spinal tract of the trigeminal nerve, whereas those of the INs were confined within the magnocellular layer of the MDH. The axonal branches of each IN formed a dense axonal mesh around the cell body of the parent neuron. Although the main bodies of the axonal fields of the INs were located in the magnocellular layer, some axonal branches extended to laminae I and II of the MDH. Immunoreactivity for NK1 receptor (substance P receptor) was found in approximately half of the PNs but not in the INs. Although no strong correlation was found between morphology and electrical membrane properties, there were some differences in electrical properties among the morphologically classified neuron groups, e.g., hyperpolarizing sag was observed in some PNs but not in the Ins; inward rectification was observed in some of the INSSs and INLAs but not in the PNs; the slow ramp depolarization and the slow afterdepolarization were observed in all INSSs examined but not in the PNs or INLAs. J. Comp. Neurol. 428: 641-655, 2000.

Morphologic features and electrical membrane properties of neurons in the substantia gelatinosa (SG) of the caudal spinal trigeminal nucleus (the medullary dorsal horn; MDH) were examined in the rat. Intracellular recording and biocytin-injection combined with histochemical staining were performed in horizontal slices. Twenty-four SG (lamina Ⅱ) neurons were recorded stably and stained successfully. Both projection neurons (PNs; n 5 9) that sent axons to regions outside the MDH and intrinsic neurons (INs; n 5 15) that sent axons only to the MDH were observed. The INs were divided into those with dense axonal arborization (INDAs; n 5 7) and those with sparse axonal arborization (INSAs; n 5 8). In the PNs, the dendrites with spines spread to all MDH layers (laminae Ⅰ-Ⅲ). The main axons sent collaterals within the SG and rostrally, caudally, or medially to laminae I and III of the MDH, interpolar spinal trigeminal nucleus, spinal tract of the trigeminal nerve, or upper cervical cord segments. In the INDAs, the dendrites arising from the rostral and caudal pole of the cell bodies mainly extended rostrally and caudally parallel to the rostrocaudal axis of the SG: the dendritic trees were elongated and oval in shape and were confined within the SG. The axonal field of each INDA, a dense mesh of axonal processes, was elongated and oval in shape and almost was confined within the SG. In the INSAs, a small, round cell body was located in the center of each dendritic field, which usually was limited within the SG. Axonal processes ran radially to spread to all layers of the MDH, constituting round or oval axonal fields. The three groups of SG neurons showed more or less different intracellular responses to current injections. In particular, adaptation of spike frequency, hyperpolarizing sag, and rebound excitation were observed in the PNs and INSAs but not in the INDAs. Slow ramp depolarization and slow afterdepolarization were recorded only in INDAs. J. Comp. Neurol. 411: 399-412, 1999.