Hermanson O, Telkov M, Geijer T, Hallbeck M, Blomqvist A
Division of Cell Biology, Faculty of Health Sciences, Linköping University, Sweden.
Eur J Neurosci. 1998 Jan;10(1):358-67. doi: 10.1046/j.1460-9568.1998.00021.x.
The dorsal lateral subnucleus of the rat pontine parabrachial nucleus is a major target for ascending nociceptive information from the spinal cord. With in situ hybridization histochemistry, using a radiolabelled cRNA probe, we demonstrate that neurones in and near the dorsal lateral subnucleus express preprodynorphin mRNA. The cRNA probe was constructed from a PCR product amplified from rat genomic DNA. Sequencing of the PCR product revealed that it corresponded to the sequence 466-1101 of the rat preprodynorphin gene exon 4. Tract tracing experiments, using injection of cholera toxin subunit B into the hypothalamic median preoptic nucleus, showed a retrograde labelling pattern of neurones in the parabrachial nucleus that was almost identical to that of the preprodynorphin mRNA expressing neurones. Double-labelling, combining immunohistochemical detection of tracer and in situ hybridization, revealed that the retrogradely labelled neurones expressed preprodynorphin mRNA. A similar double-labelling, combining in situ hybridization with immunohistochemical detection of noxious-evoked fos following formalin injection into one hindpaw of awake animals, showed that almost all fos-immunoreactive neurones in the dorsal lateral parabrachial subnucleus also expressed preprodynorphin mRNA. Quantitative analysis suggested that the evoked fos immunoreactivity was accompanied by an increased preprodynorphin mRNA expression. The findings provide evidence that neurones in the dorsal lateral subnucleus produce dynorphin and project to the median preoptic nucleus, and that noxious stimulation in awake animals synaptically activates the dynorphinergic neurones in this subnucleus. These observations are consistent with the idea of a functional and chemical heterogeneity among different parabrachial subnuclei that serves to produce specific homeostatic responses to stimuli that changes the physiological status of the organism, including tissue damage.
大鼠脑桥臂旁核背外侧亚核是脊髓上行伤害性信息的主要靶点。我们运用放射性标记的cRNA探针，通过原位杂交组织化学方法，证明背外侧亚核及其附近的神经元表达前强啡肽原mRNA。该cRNA探针由从大鼠基因组DNA扩增得到的PCR产物构建而成。对PCR产物进行测序显示，它对应于大鼠前强啡肽原基因外显子4的466 - 1101序列。通过将霍乱毒素B亚单位注射到下丘脑视前正中核进行的示踪实验表明，臂旁核中神经元的逆行标记模式与表达前强啡肽原mRNA的神经元几乎相同。将示踪剂的免疫组织化学检测与原位杂交相结合的双重标记显示，逆行标记的神经元表达前强啡肽原mRNA。在清醒动物的一只后爪注射福尔马林后，将原位杂交与有害刺激诱发的Fos免疫组织化学检测相结合的类似双重标记显示，背外侧臂旁亚核中几乎所有Fos免疫反应性神经元也表达前强啡肽原mRNA。定量分析表明，诱发的Fos免疫反应性伴随着前强啡肽原mRNA表达的增加。这些发现提供了证据，表明背外侧亚核中的神经元产生强啡肽并投射到视前正中核，并且清醒动物中的有害刺激通过突触激活该亚核中的强啡肽能神经元。这些观察结果与不同臂旁亚核之间存在功能和化学异质性的观点一致，这种异质性有助于对改变机体生理状态（包括组织损伤）的刺激产生特定的稳态反应。
