| Backgroud Spinal cord injury(SCI) is a serious nervous system trauma. At present, thereare two big difficulties in SCI recovery:(1) How to prevent nerve cell death caused by SCI, andhow to replace dead nerve cells.(2)How to inhibit local scarring, create microenvironmentsuitable to neural regeneration, promote and induce nerve growth. Traditionally, because maturenerve cells can not divide, nerve cells do not regenerate and can only be filled with glial cells andform glial scar after CNS injury in mammals. However, recent studies found that neurons ofadult animal can recover to different extents after SCI. Under adequate circumstance, someaxons of impaired neurons in CNS of mature mammals can regenerate a little, and formfunctional synapse connection with target cells. While in some immature mammals, there areaxon regeneration and function recovery. Further study suggests that neurotrophic factors canpromote neuron regeneration, thereby promote SCI recovery. Pituitary adenylatecyclase-activating polypeptide (PACAP) is a neuropeptide firstly isolated from goathypothalamus and can activate cultured rat pituitary cell adenylate cyclase, which belongs tosecretin, glucagon, vasointestinal polypeptide (VIP) and growth hormone releasing hormone(GRH) family. Recently PACAP is considered to be a new neurotrophic factor, which has somefunctions such as neurotransmitter, neuromodulator and neurotrophy. When nerve tissue isdamaged, PACAP can be excreted by peptidergic neurons, which is benefit to self-recovery ofimpaired tissue. Furthermore, as the molecular mass of PACAP is small and it can permeatethrough blood brain barrier easily, it has a latent ascendancy of administration route in treatingCNS injury.Objective To establish rat SCI models, to examine the change of PACAP expression afterSCI, and provide experimental basis for further exploring regeneration recovery mechanism afterSCI and treating SCI with neurotrophic factors.Methods Eighty four female Sprague-Dawley rats weighing 180～220g were randomlydivided into 1, 3, 7, 14, 28d after SCI and control(did no operation) groups, each group had 14rats. Rats were anesthetized with sodium pentobarbital(30mg/kg, i.p.).The median incision were at T9～11, SCI models were established with NYU weight drop device by impacting T10 segment.Materials were got at 1,3,7,14,28d after SCI, HE staining and PACAP immunochemistry(n=5)were carded out with paraffin section; Ultrastructure and localization of PACAP were detectedwith ultrathin section by transmission electron microscope(TEM) (n=3); Total RNA andprotein were extracted, the expression of PACAP mRNA and protein were examined byRT-PCR (n=3) and western blot (n=3).Results 1. Histopathology change of SCI area: Under light microscope, necrosis andcapsular space formed after hemorrhage were observed in rat's gray nucleus following SCI, theresults also showed that nerve cellular swelling, karyopyknosis in part of nerve cells, whitematter rarefaction and edema, vacuolar degeneration, some nerve fibers lysis and vanish,inflammatory cell infiltration, there was an improvement with time going.2. Immunohistochemistry observation: PACAP positive reactions were seen atcellular membrane and cytoplasm. PACAP positive expression was observed in control group,the expression of PACAP increased after SCI, with a significant increase at 7d after SCI (P＜0.05) and at 28d it was still at a high level.3. RT-PCR and western blot: There was the same tendency between PACAP mRNAand protein expression, the PACAP mRNA and protein expressed lowly in control group, thelevel increased 1d after SCI, there was an obviously increase at 7d, with a significant differencecompared with that of 3d (P＜0.05).4. TEM and immunoelectron microscopy observation: After SCI, myelin sheathdeformation and layers segregation were viewed, microfilament and microtubule in axondisordered or vanished,membrane was in bad order, free ribosome increased, the mitochondriaand rough endoplasmic reticulum swelled and vacuolar degenerated at different degrees, cristaelysed and collapsed. Colloidal gold labeling immune electron microscopy displayed that thecolloidal gold granules were found in the neuronal kytoplasm and some nerve fibers 1d afterSCI.Conclusion The increase of PACAP expression after SCI indicated that endogenousPACAP may promote the survival and regeneration of injured spinal neurons. As a neurotrophicfactor, PACAP may have some relations with self-recovery after SCI.
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