Immunohistochemical, fluorescence and a number of biochemical techniques have demonstrated that high concentrations of cholecystokinin (CCK), somatostatin (SS), substance P (SP), vasoactive intestinal polypeptide (VIP), L-glutamate, 5-hydroxytryptamine (5-HT) and norepinephrine (NE) are present in the superficial parts of the spinal dorsal horn, the same area where small myelinated (A(delta)) and unmyelinated (C) primary afferent fibers are known to terminate. The intent of this project was to study the actions of these substances on the excitability of functionally identified dorsal horn neurons and single cutaneous primary afferent fibers in the cat spinal cord in vivo;Applied iontophoretically and/or by pressure microinjection, CCK-8 and VIP caused a moderate to strong excitation of a majority of tested neurons located in laminae I-VII of intact cat spinal cord. The excitation was not limited to a single population of neurons but was observed in all categories of units recognized in spinal preparations of cats in this area on the basis of their excitability by different kinds of cutaneous afferent input. These results are consistent with the possibility that CCK-8 and VIP act in the dorsal horn of the spinal cord as neurotransmitters or neuromodulators. While in intact cats SP excited about 31% of units activated by innocuous stimulation of the skin and about 80% of units activated by noxious stimulation, this peptide caused excitation of all spinal cord neurons tested in p-chlorophenylalanine (p-CPA)-pretreated cats. Thus, irrespective of their excitatory input, the sensitivity of spinal neurons to SP appears to be increased in the p-CPA-pretreated cats;In order to determine more directly whether SS, L-glutamate and NE exert presynaptic action on the primary afferent fibers, we have measured the electrical excitability changes of intraspinal single cutaneous primary afferent C- and A-fibers during their iontophoretic and/or micropressure application into the spinal cord. SS and L-glutamate increased excitability in about 50% of all fibers tested. NE decreased excitability in a majority of fibers tested. In addition, we have found that both (alpha)- and (beta)-receptor agonists cause a reduction of the excitability of C- and A-fibers, clonidine being the most potent in this respect and isoproterenol the least potent. These results indicate that SS, L-glutamate and NE, in addition to previously shown postsynaptic actions, also modify excitability of intraspinal cutaneous primary afferent fibers.