The pontine parabrachial nucleus (PB) is a major target for ascending fibers from nociresponsive dorsal horn neurons. Several different neuropeptides have been identified in the PB. By using double-labeling methods that combine in situ hybridization histochemistry with retrograde tract tracing and immunohistochemistry, we have examined the subnuclear localization of preprocholecystokinin mRNA (ppCCK)-containing neurons, investigated their efferent projection, and analyzed their expression of intracellular signaling substances that may be of importance for nociceptive processing. The results show that neurons containing ppCCK are preferentially localized to the superior lateral subnucleus (PBsl), whereas other subnuclei, such as the dorsal lateral, external lateral, central lateral, and ventral lateral subnuclei, and the Kölliker-Fuse nucleus, contain only moderate to small numbers of such neurons. Injections of the retrograde tracer cholera toxin subunit b into the ventromedial hypothalamus demonstrated that ppCCK-containing neurons in PBsl were projection neurons. Following nociceptive stimulation, the ppCCK-containing neurons expressed FOS protein as well as phosphorylated cyclic AMP-responsive element-binding protein (CREB). In addition, Ca2+/calmodulin-dependent kinase II (CaMKII) was heavily and rather selectively expressed in PBsl and was co-localized to ppCCK-containing neurons. These observations show that nociceptive stimuli activate a cholecystokinin pathway from the parabrachial nucleus to the ventromedial hypothalamus that may be important for homeostatic responses to tissue damage, and point to a putative intracellular route for Ca2+-mediated FOS transcription via CaMKII and CREB for the regulation of ppCCK transcription.