Although the molecular and cellular responses to injury in the central nervous system (CNS) have been widely investigated, few studies have examined the potential variations between direct and indirect neuronal injury. To differentiate between the response to axotomy and deafferentation, two central cholinergic populations were analyzed: the horizontal limb of the diagonal band of Broca (HLDB) and the interneurons in the corpus striatum (CS). At time points from one hour to eight weeks postinjury the levels of choline acetyltransferase (ChAT) mRNA and protein were assessed by in situ hybridization and immunohistochemistry. Also examined was the expression of the immediate early gene product, c-fos. One week post axotomy, neurons in the HLDB exhibited an increase in the levels of ChAT mRNA without a concomitant increase in ChAT protein, followed by a steady decrease reaching a nadir in both parameters at eight weeks. In contrast, a transient increase occurred at one week postdeafferentation in the levels of both ChAT mRNA and protein in the interneurons of the CS. Axotomized neurons in the HLDB did not exhibit either c-fos mRNA or protein expression, while robust fos induction occurred after one hour in deafferented neurons in the CS. These data demonstrate that the molecular and cellular responses differ following direct and indirect neuronal injury. Furthermore, they suggest that in these central cholinergic populations deafferentation may result in cellular hyperactivity and cell survival while axotomy results in decreased cellular activity and subsequent cellular regression.