Previous work has shown that the inbred Lewis and Fischer 344 rat strains differ in several behavioral measures related to mesolimbic dopamine function. Moreover, the ventral tegmental area (VTA) of the Lewis rat has been shown to contain higher levels of tyrosine hydroxylase compared to that of the Fischer rat by blot immunolabeling procedures. To investigate structural correlates of this biochemical difference, an immunohistochemical study of VTA dopaminergic neurons in these two strains was undertaken. Results show that the density and total number of tyrosine hydroxylase-positive neurons in the VT 4 of the Lewis rat is about 50% of that found in the Fischer rat. In contrast, examination of the substantia nigra in the same sections revealed no differences in the density and number of tyrosine hydroxylase-positive cells between these strains. Fischer-Lewis strain differences were also evident for cholecystokinin immunoreactivity in the VTA, with much lower levels seen in the Lewis rat, consistent with the known colocalization of this neuropeptide in many VTA dopamine neurons. The finding of 50% fewer tyrosine hydroxylase-positive neurons in the VTA of the Lewis rat, along with our earlier results showing 45% higher levels of tyrosine hydroxylase by blot immunolabeling, would suggest much higher levels of tyrosine hydroxylase per VTA neuron in this strain. However, no obvious strain difference in the cellular intensity of tyrosine hydroxylase immunoreactivity could be detected by immunohistochemistry. Finally, the density of VTA dopamine neurons was assessed in 1-week-old Fischer and Lewis rats. In contrast to the results obtained for adult animals, no difference in the number of tyrosine hydroxylase-positive neurons was apparent in these young animals, indicating that the Fischer-Lewis strain difference in VTA dopamine neurons appears later in postnatal development. These anatomical findings shed new light on the differences in the mesolimbic dopamine system between Fischer and Lewis rats that may contribute to the behavioral differences exhibited by these animals.