Brain 12-HETE formation in different species, brain regions, and in brain microvessels.

We used gas chromatography/mass spectrometry to measure brain 12-HETE (12-Hydroxy-5,8,10,14-eicosatetraenoic acid) formation from endogenous arachidonic acid in different species and different brain regions and in isolated brain microvessels. When blood-free brain slices were incubated for 20 minutes we found that the rabbit and cat brain incubates contained little 12-HETE when compared to rat and mouse brain incubates. Further in vitro studies of various rat brain regions showed a generally even distribution of 12-HETE. When isolated rat or rabbit microvessels were incubated and analyzed, we found 1 and 0.25 micrograms, respectively, of 12-HETE/mg of microvessel protein. Also, rabbit brain had limited or no capacity to actively metabolize tritiated 12-HETE. In summary, these studies show substantial species variation with respect to brain formation of 12-HETE and indicate that the vasculature is a potentially significant contributor to the 12-HETE found in whole brain tissue.

Inhibition of bradykinin- and kallikrein-induced cerebral arteriolar dilation by a specific bradykinin antagonist.

We have previously shown that topical brain application of kallikrein, an enzyme which converts kininogen to bradykinin, induces rabbit pial arteriole dilation. The purpose of the present investigation was to utilize a newly developed competitive kinin receptor antagonist to test the hypothesis that kallikrein-induced dilation was due to the conversion of brain kininogen to vasoactive kinins. As in our previous study, we measured rabbit pial arteriole diameter with a microscope using the closed cranial window technique. The kinin antagonist (6 microM) reduced the dose-dependent dilation produced by bradykinin and blocked the dilation induced by kallikrein. In addition, the kinin antagonist was specific since it did not alter the cerebral arteriole dilation produced by adenosine, acetylcholine, or vasoactive intestinal polypeptide. These experiments provide further evidence for a possible role of the endogenous brain kallikrein-kinin system in the modulation of the cerebral circulation and provide the necessary pharmacologic foundation for future use of this antagonist in testing the role of kinins in the normal or altered cerebral circulation.

The effects of catalase, indomethacin and FPL 55712 on vascular permeability in the hamster cheek pouch following scald injury.

The effects of scald injury and scald injury with pretreatment on plasma and water content were studied in the superfused hamster cheek pouch. Catalase (30,000 Units/kg), indomethacin (2.5 mg/kg) and FPL 55712 (2 mg/kg) were administered prior to 10 second scald with 100 degrees C normal saline. Plasma content was measured with 125 I serum albumin and water content was calculated by loss on drying. Significant (P less than 0.05) decreases in plasma content compared to scald alone were observed following pretreatment with catalase (29%) indomethacin (31%) or FPL 55712 (52%). Water content in scalded pouches was significantly reduced by catalase (29%) and FPL 55712 (42%). Pretreatment when combined also resulted in significant reduction of plasma content but not water content. Results suggest that free-radicals, prostaglandins and leukotrienes are involved in vascular response by scald injury.

A modified technique for permeability studies in the superfused hamster cheek pouch.

We report here a modification of the superfused hamster cheek model for use in vascular permeability studies. Radio-iodine labeled serum albumin (I-125 RISA) is injected prior to the superfusion period. Plasma content is calculated on a microliter/100 mg wet weight basis and compared to the contralateral (non-superfused) cheek pouch. Water content is calculated on a percentage basis and compared in the same manner. Results demonstrate that superfusion causes an increase in permeability of protein and water. Plasma content is reduced by catalase, indomethacin or FPL 55712 pretreatment, suggesting that free-radicals, prostaglandins and leukotrienes are released during superfusion. Water content increase is refractory to pretreatment. The advantages of this system and its application are discussed.

Free-radical inhibition of ATPase in hamster cheek pouch homogenates.

The effects of free-radicals generated by either the oxidation of hypoxanthine by xanthine oxidase (HX/XO) or the lipoxidation of arachidonic acid (AA) on the ATPase of the hamster cheek pouch has been studied. Cheek pouches were removed from female golden syrian hamsters and homogenized. ATPase activity was measured by the production of Pi at 37 degrees. HX/XO and AA were added at a final concentration of 9.6 X 10(-5) M HX with 5 X 10(-2) units HX and 5 X 10(-5) M AA with and without 1 X 10(-4) M ouabain. HX/XO produced a 24.7% inhibition alone and 35.0% when combined with ouabain. Ouabain alone produced a 7.1% inhibition. AA produced a 23.6% inhibition alone and 24.3% inhibition when combined with ouabain. Ouabain alone produced a 5.4% inhibition in this series. When AA was added in doses ranging from 1 X 10(-5) to 2 X 10(-3) M, a plot of percent inhibition versus log dose followed a typical sigmoid type curve. The IC50 was 1.5 X 10(-4) M. These results suggest that free-radicals are capable of inhibiting the ATPase found in the hamster cheek pouch tissues. The possible modes of action of the free-radicals in producing this inhibition are discussed.