The Expression Of ANB In The Lumbosacral Spinal Cord Of Aged Rat And Its Related Features

The lumbosacral spina cord is important for controlling the function ofthe large intestine,the pelvicmuscel and the reproductive organs. During agingprocess, the spinal lumbosacral segments show aging-related changesincluding physical and functional changes. Nitric oxide (NO) is one of neuralinformation materials in the signal transduction, which plays an important rolein the peripheral and central nervous system. It is suggested that theabnormalities of pelvic organ function might be associated with the alteredNO expression in lumbosacral spinal cord. During the aging process, there arethe expression changes of NO and the appearance of age-related nicotinamideadenine dinucleotide phosphate-diaphorase positive bodies (ANB) in thelumbosacral spinal cord. Sex hormone (testosterone) undergoes a progressivedecline during normal male aging. The relationship between the appearance ofANB and the reduction of testosterone in the aging male is worthwhile to bestudied.5-hydroxytryptamine (5-HT) and dopamine (DA) neural systems playimportant roles in the control of micturition reflex in the lumbosacral spinalcord. It is found that both5-HTergic and DAergic innervation was decreasedgradually during the aging process. Because till now, other neurochemicalidentity of ANB is still not known except NO,whether coexistence of ANBwith5-HT or DA that are related most with the control of micturition reflexshould be investigated.Objective: To observe the expression of ANB in the lumbosacral spinalcord of aged rats as well as the effect of testosterone propionate on it, and toclarify5-HTergic or DAergic identity of ANB.Motheds: Male Wistar rats were randomly divided into6months (6Mon),21months (21Mon) and24months (24Mon) group.Testosterone propionate (TP) was administered either by subcutaneous injection or intranasal delivery to21Mon (21Mon-sc.TP or21Mon-int.TP) and24Mon rats (24Mon-sc.TP or24Mon-int.TP). The same treatment usingsesame oil as a vehicle to21Mon (21Mon-sc.con or21Mon-int.con) and24Mon (24Mon-sc.con or24Mon-int.con) rats as control. Testosteronepropionate was given at a dose of2.0mg/kg, at5:00PM to6:00PM, once aday from the18-month old to21-month old or from21-month old to24-monthold of rats.All the rats were perfused transcardially with a fixative containing4%paraformaldehyde. The lumbosacral segments were cryoprotected by30%sucrose solution and then cut using cryomicrotome. Corresponding spinalsections were respectively collected for NADPH-d histochemistry andNADPH-d histochemistry combining with5-HT or tyrosine hydroxylase (TH)immunohistochemical double labelling in order to observe the appearance ofANB in the lumbosacral cord of aged rats and the effect of TP on it, as well asrelationship the ANB with5-HT or DA.Result:1.NADPH-d histochemistry(1)The general observation of the lumbosacral sections: NOS positiveneurons in the section of lumbosacral spinal cord were stained blue and theyappeared golgi-like. NOS neurons were medium-sized, dark-or light-stained,spindle-shaped, triangular or rounded. they have2-4processes and the nucleuswas not stained. The ANB were detected in the SDH, LT, ILN, DCN and CCof the lumbosacral spinal cord. They were highly-dense spheroidal or irregularshapes with sharp edges.(2)The number change of ANB in the lumbosacral sections: There was noANB observed in the lumbosacral sections of6Mon group, while ANBs couldbe detected in the lunbosacral sections of21Mon and24Mon group of rats.ANBs were much more in24Mon group than in21Mon group. ANB numberin24Mon-int.con group was significantly40%(P<0.05),65%(P<0.05),69%(P<0.05),88%(P<0.05)and39%(P<0.05) more than that in21Mon-int.congroup in SDH, LT, ILN, DCN and CC respectively. (3)The influence of TP upon the ANB:①Subcutaneous injection of TP:Subcutaneous injection of TP could reduce ANB numbers in21Mon-sc.TP rats.The number of ANB was significantly40%(P<0.05),34%(P<0.05),40%(P<0.05),60%(P<0.05)and82%(P<0.05) less in21Mon-sc.TP than that in21Mon-sc.con in SDH, LT, ILN, DCN and CC respectively.The number ofANB was5%,13%,3%and6%less in24Mon-sc.TP than that in24Mon-sc.con in SDH, LT, ILN and DCN respectively, but the difference didnot reach significant level.②Intranasal administration of TP: The number ofANB was significantly20%(P<0.05),22%(P<0.05),32%(P<0.05),38%(P<0.05)and44%(P<0.05) less in21Mon-int.TP than that in21Mon-int.con in SDH, LT,ILN, DCN and CC respectively. The number of ANB was6%,10%and10%less in24Mon-int.TP than that in24Mon-int.con in ILN, DCN and CCrespectively, but the difference did not reach significant level.(4)The comparison of efficacy of intranasal delivery of TP withsubcutaneous injection of TP on the ANB: There was no significant differencein the effect of TP on ANB number between21Mon-sc.TP and21Mon-int.TP,which suggest that subcutaneous injection and intranasal administration of TPhas same efficacy in reduction of ANB number.2.NADPH-d histochemistry conbining with5-HT or THimmunohistochemistry: There was no coexistence of ANB with5-HT or THobserved in lumbosacral sections both in21-month and24-month old rats.Conclusion:1. The occurrence of ANB in lumbosacral spinal cord isrelated to aging. ANB increased gradually during the aging process.2. ANBappears to be related to low testosterone in male aging. The supplement oftestosterone propionate can reduce ANB number in aged male.3.Supplementof testosterone propionate by subcutaneous injection or intranasaladministration has same efficacy in reduction of ANB number.4. ANB doesnot coexist with either5-HT or DA.