Nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) histochemistry was used to study the morphology and development of neurons that metabolize nitric oxide (NO) in the frontal cortex of human fetuses aged from 13 weeks of gestation (13W) to term, to investigate whether the two distinct types of NO neuron described in the adult develop differently. Large, heavily stained, sparsely spiny, non-pyramidal neurons (Type I) develop by 15W mainly in the subplate (SP) of the cortical Anlage. They achieve an adult-like pattern by 32W, distributed thoughout the cortex and subcortical white matter, but with the highest concentration in the white matter. Small, lightly stained cells (Type II) develop later (32W) thoughout the cortex, but especially in layers II-IV. and increase in number to term. NADPH-d-positive dendrites and axons appear in the cortex and white matter by 15W. They include thick, radially oriented, dendritic processes from Type I neurons in SP and CP Their arbors expand and mature between 17 and 28W. Fine horizontal axons are visible in layer I by 17W. Others develop in layers II-IV from 28W, and have reached a high degree of development by term. NADPH-d-positive axons in the cortex seem to have both intrinsic and extrinsic origins. Thus the two types of NADPH-d neurons found in adult primate, including human, cortex are reflected by different developmental forms prenatally. It is concluded that NO-metabolizing neurons in the human cortex may be involved in various aspects of development, including morphological and functional maturation, and that the late-developing Type II neurons may represent a cell line specific to primates, perhaps related to the development of their higher cortical activity and of potential importance in the pathophysiology of diseases of cognitive function.