Artifactual dendritic beading in rat spinal cord induced by perfusion with cold saline and paraformaldehyde.
Extensive dendritic beading of MAP2 (microtubule-associated protein 2) immunoreactivity has previously been observed in the contused rat spinal cord. However, we have also observed dendritic beading in occasional uninjured animals. The purpose of this study was to examine the possibility that perfusion conditions contributed to the dendritic beading. Under deep anesthesia, uninjured rats (adult female Long-Evans, 200-225 g) were transcardially perfused with 0.9% saline solution followed by 4% paraformaldehyde at cold (4 degrees C) or warm (20 degrees C) temperature, and at a low (20 ml/min) or high (50 ml/min) flow rate. Dendrites were visualized by MAP2 immunoreactivity. The results demonstrate that perfusion with cold solutions at a high flow rate induces pronounced dendritic beading, and when perfused at a low flow rate, results in moderate dendritic beading. Warm perfusates did not induce dendritic beading when administered at a low flow rate, but occasional beading was observed with a high flow rate. Western blots revealed spectrin breakdown, but not MAP2 loss, in rats perfused with cold saline solution at a high flow rate, conditions that also resulted in dendritic beading. These findings demonstrate that dendritic morphology is sensitive to both temperature and flow rate of the perfusate. Warm fixative and a low perfusion flow rate minimized the perfusion-induced dendritic beading.
Zhang
Shu-Xin
SX
Spinal Cord and Brain Injury Research Center, University of Kentucky College of Medicine, Lexington, KY 40536, USA. szhang@scs.cbeyond.com
Holmberg
Eric G
EG
Geddes
James W
JW
D000704:Analysis of Variance; D000818:Animals; D003080:Cold Temperature / Q000009:adverse effects; D003712:Dendrites / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D005260:Female; D005404:Fixatives / Q000494:pharmacology*; D005557:Formaldehyde / Q000494:pharmacology*; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D010477:Perfusion / Q000009:adverse effects / Q000379:methods; D011108:Polymers / Q000494:pharmacology*; D051381:Rats; D020318:Rats, Long-Evans; D012965:Sodium Chloride / Q000494:pharmacology*; D013049:Spectrin / Q000378:metabolism; D013119:Spinal Cord Injuries / Q000473:pathology*
D005404:Fixatives; C489787:MAP2 protein, rat; D008869:Microtubule-Associated Proteins; D011108:Polymers; D013049:Spectrin; D005557:Formaldehyde; D012965:Sodium Chloride; C003043:paraform