Application of morphine prior to noxious stimulation differentially modulates expression of Fos, Jun and Krox-24 proteins in rat spinal cord neurons

The expression of Fos, Jun and Krox-24 proteins was investigated in spinal cord neurons of the rat 2, 4 and 8 h following noxious thermal stimulation of one hind-paw and pre-treatment with morphine. The number of neurons expressing c-Fos, c-Jun, Jun B and Krox-24 were maximal after 2 h and thereafter declined. The number of Fos B and Jun D immunoreactive neurons increased constantly for up to 8 h with Jun D showing expression above baseline only after 4 h following stimulation. Intravenous application of morphine (5 and 10 mg/kg) 20 min before noxious heat stimulation decreased the expression of all six proteins at any time-point with a predilective effect on neurons of deeper laminae of the dorsal horn. The suppressive effects of morphine were more pronounced with the higher dose of morphine and completely reversed by intravenous naloxone (1 and 10 mg/kg). The temporospatial patterns of expression following morphine were similar to those seen without morphine, but in a much smaller number of neurons and with a shorter time-course. However, despite the high dose of morphine and continuous halothane anaesthesia during the whole experimental procedures, a considerable number of neurons expressing the various genes remained in all laminae of the spinal cord. At 2 h following noxious heat stimulation morphine had decreased the number of labelled neurons for c-Fos, Fos B, Krox-24, c-Jun and Jun B to 30-60% of control levels in laminae I-II and to 10-30% in laminae III-VII,X of the spinal cord. At 4 h the level of reduction had further increased while Jun D was only moderately reduced to 75% in all laminae of the spinal cord. Eight hours following noxious heat plus morphine application we did not detect noxious evoked immunoreactivity for c-Fos, Krox-24, c-Jun and Jun B, while there was residual labelling for Fos B in the superficial dorsal horn and for Jun D in laminae I-VII and X of the spinal cord. The different temporospatial pattern of immediate early gene expression in neurons of the spinal cord dorsal horn following noxious stimulation suggest that variable transcription complexes may interact with DNA regulatory sequences and could thus activate alternative secondary response genes, even under protection of a high dosage of morphine applied before noxious stimulation.