In this work, the time course of appearance, distribution and morphology of diaphorase-positive neurons were studied in the developing cingulate cortex of the human brain during the second half of gestation. Five human fetuses at 18, 20, 25, 30 and 35 weeks postovulatory (wpo) were examined. The brain tissue was reacted by an indirect histochemistry protocol for detection of NADPH-diaphorase activity. Labeled neurons were identified at the microscope and documented photographically or by computer-aided charts. We have found that heavily labeled neurons (type I) first appear in the subplate (SP) between 20 and 25 wpo, and in the cortical plate (CP) between 25 and 35 wpo. By 35 wpo, CP neurons were both type I and type II (lightly labeled neurons). In addition, we observed 4 different morphological types among subplate neurons, very similar to callosally-projecting subplate cells (as described previously by our group). We concluded that medial nitridergic neurons of humans appear prenatally according to the usual gradient of cortical maturation -- first in the subplate and later in the cortical plate. Also, we suggest that some of the diaphorase-positive neurons in the transient subplate could possibly be callosal.