Birth-date dependent arrangement of spinal enkephalinergic neurons: evidence from the preproenkephalin-green fluorescent protein transgenic mice

Enkephalin (ENK) has been postulated to play important roles in modulating nociceptive transmission, and it has been proved that ENKergic neurons acted as a critical component of sensory circuit in the adult spinal cord. Revealing the developmental characteristics of spinal ENKergic neurons will be helpful for understanding the formation and alteration of the sensory circuit under pain status. However, the relationship between the embryonic birth date and the adult distribution of ENKergic neurons has remained largely unknown due to the difficulties in visualizing the ENKergic neurons clearly. Taking advantage of the preproenkephalin-green fluorescent protein (PPE-GFP) transgenic mice in identifying ENKergic neurons, we performed the current birth-dating study and examined the spinal ENKergic neurogenesis. The ENKergic neurons born on different developmental stages and their final location during adulthood were investigated by combining bromodeoxyuridine (BrdU) incorporation and GFP labeling. The spinal ENKergic neurogenesis was restricted at E9.5 to E14.5, and fitted in the same pattern of spinal neurogenesis. Further comparative analysis revealed that spinal ENKergic neurons underwent heterogeneous characteristics. Our study also indicated that the laminar arrangement of ENKergic neurons in the superficial spinal dorsal horn depended on the neurogenesis stages. Taken together, the present study suggested that the birth date of ENKergic neurons is one determinant for their arrangement and function.