Three new cholinergic markers were employed to study the cholinergic innervation in the thalamus of adult macaque monkeys. They were: two antibodies against choline acetyltransferase (ChAT), one polyclonal and one monoclonal; and a polyclonal antibody against the vesicular transporter of acetylcholine (VAChT), a powerful new marker that colocalizes with ChAT. This approach led to an unexpected finding: the three antibodies positively immunostained a population of neurons in the paracentral nucleus. The immunostained cells are confined to the dorsal region of this nucleus along its rostrocaudal extent. Measurement of the somatic areas of the immunostained neurons indicated that they correspond to a population of large neurons thought to be projection neurons. Because dorsal paracentral neurons are known to project to the dorsal striatum and specific cortical areas involved in visual and visuomotor mechanisms, these structures might be modulated by cholinergic thalamic neurons.