During lactation, suckling stimulus causes various behavioral and endocrinal changes, such as the maternal behavior, increase in food intake, and suppression of reproductive function. Previous experiments have shown that suckling stimulus is conveyed to the brain stem via the spinal cord and then to the hypothalamus followed by the various brain regions. In the present study, we purposed the identification of neurons conveying suckling stimulus from the brain stem to the hypothalamus in lactating rats. We investigated lactation-associated factors in the brain stem using the quantitative RT-PCR analysis. We found significant increase in the expression of calcitonin gene-related peptide (CGRP) alpha in the brain stem during lactation. Next, we examined distribution of the CGRP neural cell bodies and fibers in lactating rats by immunohistochemistry. Many CGRP-immunoreactive (ir) neural cell bodies were found in the peripeduncular nucleus of the mesencephalon (PP) and posterior intralaminar nucleus of the thalamus (PIL), and CGRP-ir fibers were found in the ventromedial nucleus of the hypothalamus and caudal part of caudate-putamen. The ratio of cFos-ir in the CGRP-ir neurons in the PP and PIL in suckling-stimulated mother rats was greater than that in non-stimulated control rats. These results suggested that CGRP neurons in the PP and PIL play a key role in conveying suckling stimulus to the hypothalamus during lactation.