Repeated, short-term ischemia augments bradykinin-mediated opening of the blood-tumor barrier in rats with RG2 glioma

The objective of this study was to investigate the effects of repeated, sho rt-term ischemia on bradykinin-mediated permeability of the blood-brain bar rier (BBB) and the blood-tumor barrier (BTB). The mechanism by which bradyk inin transiently opens the BTB, involves B2 receptors, Ca2+ flux, nitric ox ide (NO) and cyclic CMP (cGMP). Since global and focal cerebral ischemia ar e known to increase levels of brain nitric oxide synthase bNOS) and endothe lial nitric oxide synthase (eNOS) we tested the hypothesis that bradykinin may increase the BTB permeability to a greater extent under,ischemic rather than nonischemic conditions. The vertebral arteries in female Wistar rats were coagulated immediately after intracerebral implantation of RG2 glioma. Short-term ischemia was produced in some rats by a modification of the fou r-vessel occlusion procedure for incomplete forebrain ischemia, in which th e common carotid arteries were clamped daily for 15 min on days 7, 8 and 9 after tumor implantation, after which reperfusion was allowed. On day 10 af ter tumor implantation, bradykinin (10 pg kg(-1) min(-1)) or phosphate-buff ered saline (PBS) was infused for 15 min into the right carotid artery of a nesthetized, sham-operated (nonischemic controls) and ischemic rats, follow ed by an intravenous bolus (100 mu Ci kg(-1)) each of [C-14]-iodo-antipyrin e (IAp), [C-14]-dextran or [C-14]-aminoisobutyric acid (AIB) to measure reg ional cerebral blood flow (rCBF), blood volume, or unidirectional transfer constant Ki, respectively, by quantitative autoradiography. A single 15-min ischemic episode significantly decreased rCBF in the tumor center (158.9 /- 17.33 in control vs. 58.78 +/- 24.45 m/100 g(-1) min(-1) in ischemic gro up; p < 0.01) and in the tumor periphery (106.82 +/- 7.34 in control vs. 70 .55 +/- 26.66 ml 100 g(-1) min(-1) in ischemic group; p < 0.05), Respective mean blood volume in tumors (11.7 +/- 13.3, 12.7 +/- 14.0, and 13.3 +/- 14 .5 mu lg(-1)) from ischemic-PBS nonischemic-bradykinin, and ischemic-bradyk inin groups, respectively, was not significantly different; mean blood volu me in normal brain (3.7, 3.1 and 3.8 mu lg(-1)) was not significantly diffe rent among these groups either. Intracarotid infusion of bradykinin followi ng repeated ischemia significantly increased mean Ki, as compared to bradyk inin infusion in nonischemic controls, in both the tumor center (36.60 +/- 8.4 vs. 22.90 +/- 4.61 mu lg(-1) min(-1), p < 0.05) and in tumor periphery (17.70 +/- 5.93 vs. 8.50 +/- 4.42 mu lg(-1) min(-1), p < 0.05). Mean Ki val ues for tumor center and tumor periphery of ischemic rats receiving intraca rotid bradykinin were 3-fold greater than those of nonischemic rats infused with PBS. Immunohistochemical and Western blot analyses showed that repeat ed, short-term ischemia significantly increased the levels of bNOS in tumor cells and eNOS in tumor capillaries, but neither induced iNOS nor affected B2 receptor levels in tumor cells in vivo, as compared with nonischemic co ntrols. Taken together, these results demonstrate for the first time that r epeated, short-term ischemia augments bradykinin-mediated opening of the BT B. We conclude that the elevated intratumoral levels of bNOS and eNOS may ' prime' the NO generating capacity of tumor cells. Consequently, increased d e novo synthesis and a correspondingly elevated concentration of NO within the tumor, therefore, may be one mechanistic explanation for the significan tly increased, bradykinin-mediated BTB opening under ischemic conditions, r eported here.