The effect of the adrenocorticotropin-(4-9) analogue, ORG 2766, and of dizolcipine (MK-801) on infarct volume in rat brain.

The purpose of this study was to evaluate whether the synthetic adrenocorticotropin-(4-9) (ACTH-(4-9)) analogue ORG 2766, HMet(O2)-Glu-His-Phe-D-Lys-Phe-OH, which has been shown to have beneficial effects on both the recovery from experimentally induced lesions of the central nervous system and peripheral nerve degeneration, has a protective effect on focal ischemic neuronal damage. The NMDA receptor antagonist dizolcipine (MK-801), a very potent neuroprotective drug, was used as positive reference compound. Isoflurane-anesthetized rats had the middle cerebral artery occluded using either an intravasal or an extravasal technique, because pilot experiments had shown differences in the severity of ischemia for the two middle cerebral artery occlusion techniques. MK-801, 500 microg kg(-1) min(-1), or saline was administered i.v. 30 min after occlusion of the middle cerebral artery. In the ACTH-(4-9) analogue/saline group, 10 and 150 microg/kg of the analogue, or saline was injected s.c. both directly after and 24 h after occlusion. The ACTH-(4-9) analogue treatment had no effect on the infarction volume in either model of middle cerebral artery occlusion, whereas MK-801 caused a significant reduction in the volume of cortical infarction in both models. We conclude that, although ORG 2766 is known to enhance the recovery from experimentally induced lesions of the central nervous system through a neurotrophic action and has proven to have significant beneficial effects on peripheral nerve regeneration, it did not prevent ischemic neuronal damage after intravasal or extravasal middle cerebral artery occlusion in rats. The results with MK-801, which caused significant reductions in the volume of cortical infarction in both models of middle cerebral artery occlusion, with clearly the largest reduction in the intravasal middle cerebral artery occlusion model, again indicate that there are differences in the severity of the cerebral ischemia which the two models produce in the rat brain.

Collateral hemodynamics after middle cerebral artery occlusion in Wistar and Fischer-344 rats.

We investigated whether the difference in infarction volume after occlusion of a long proximal segment of the middle cerebral artery between Wistar and Fischer-344 rats, is caused by differences in collateral blood flow rate through leptomeningeal anastomoses. In view of the retrograde direction of collateral blood flow into the middle cerebral artery territory, we developed parasagittal laser-Doppler flowmetry. Using this method two laser-Doppler probes are placed on the cerebral cortex: probe 1 is placed near the anastomoses between the middle- and anterior cerebral artery, probe 2 is placed 2 mm further away from these anastomoses than probe 1. We found in both rat strains a comparable relation between the areas under the curve of the signal measured by both laser-Doppler probes for 2 h after middle cerebral artery occlusion. This relation is considered to be a measurement of the collateral blood flow rate into the middle cerebral artery territory through leptomeningeal anastomoses after middle cerebral artery occlusion. We conclude that collateral blood flow for the two strains were essentially similar for the initial 2 h after MCA occlusion. Although these collateral blood flows could have been different at a later time, it is unlikely that the interstrain difference in cerebral infarction volume between Wistar and Fischer-344 rats after proximal middle cerebral artery occlusion is caused by an apparent interstrain difference in the magnitude of collateral blood flow rate through leptomeningeal anastomoses. The parasagittal laser-Doppler flowmetry technique we developed for these experiments is currently successfully used in our laboratory to evaluate the efficacy of hemodynamically active pharmacotherapeutical agents in raising the collateral blood flow rate into the middle cerebral artery territory after middle cerebral artery occlusion.

Rat middle cerebral artery occlusion by an intraluminal thread compromises collateral blood flow.

We compared in Wistar rats collateral blood flow through leptomeningeal anastomoses after middle cerebral artery occlusion using craniotomy ('extravasal occlusion'), which results in a small volume of cerebral infarction, and after intraluminal thread occlusion ('intravasal occlusion'), which produces a large volume of cerebral infarction. Simultaneous laser-Doppler flowmetry with two probes placed on the cerebral cortex was used to measure and compare collateral blood flow. Extravasal occlusion caused a cortical blood flow reduction along a gradient in lateral direction, whereas blood flow reduction after intravasal occlusion was more uniformly distributed. It is concluded that permanent intravasal occlusion compromises collateral blood flow and therefore may not be a suitable model for measuring the ability of pharmacotherapeutic agents, if any, to improve collateral blood flow acutely after middle cerebral artery occlusion. The two models can be useful for testing drugs on parenchymal neuroprotective properties. Thereby, the intraluminal technique is preferred because of the possibility to study reperfusion damage when transient occlusion is applied.

The effects of gamma 2-melanocyte-stimulating hormone and nimodipine on cortical blood flow and infarction volume in two rat models of middle cerebral artery occlusion.

We observed that the pro-opiomelanocortin-derived neuropeptide, gamma 2-melanocyte-stimulating hormone (gamma 2-MSH), has various peripheral and central hemodynamic effects in the rat, including a marked enhancing effect on cerebral blood flow. This hemodynamic profile might be of interest in the pharmacotherapeutic approach to acute cerebral ischemia. Being an adrenocorticotropin (ACTH) analogue, gamma 2-MSH might also possess direct neuronal protective properties. Therefore, in two rat models of focal cerebral ischemia we studied the effects of gamma 2-MSH, with nimodipine, a Ca2+ channel antagonist, as a reference compound, on parasagittal laser-Doppler-assessed cortical blood flow and infarction volume. In isoflurane-anesthetized Wistar and F344 rats i.v. bolus infusions (four in total) of gamma 2-MSH or nimodipine or their vehicle controls were given 1 h before, 1 min after, and 1 h and 2 h after occlusion of the middle cerebral artery. We used both an intravasal and an extravasal middle cerebral artery occlusion technique because pilot experiments had shown differences in the severity of ischemia with the two techniques. gamma 2-MSH (100 nmol/kg in 1 min) increased cortical blood flow significantly but transiently, both pre- and post-ischemically, whereas nimodipine (20 micrograms/kg in 1 min) increased cortical blood flow only pre-ischemically in both models of middle cerebral artery occlusion. gamma 2-MSH had no effect on cortical and striatal infarction volume, while nimodipine caused a significant reduction of cortical infarction volume in the extravasal middle cerebral artery occlusion model. To conclude, despite its hemodynamic and possible neuroprotective properties, gamma 2-MSH did not prevent ischemic neuronal damage after middle cerebral artery occlusion in rats. This might be partly due to the short half-life of the peptide, leading to a transient increase in cortical blood flow and short neuronal exposure time, suggesting that prolonged infusion of the neuropeptide might be required. The results with nimodipine support the notion that it attenuates cortical ischemic damage, independently of effects on cerebral hemodynamics.

Differences in striatal extracellular amino acid concentrations between Wistar and Fischer 344 rats after middle cerebral artery occlusion.

We hypothesized that the interstrain difference between Wistar and Fischer-344 (F344) rats in cerebral infarction volume after proximal middle cerebral artery (MCA) occlusion might be explained by differences in excitotoxicity between both rat strains. Using microdialysis we measured during a 5 h period after MCA occlusion the release of aspartate, glutamate and taurine in the cerebral cortex and the striatum. The volume of striatal infarction was comparable in Wistar and F344 rats. We found, however, in Wistar rats a significantly higher striatal efflux of aspartate and glutamate than in F344 rats, whereas the striatal taurine efflux was of a similar magnitude in the two strains. Because of the (variably) smaller volume of cortical infarction in Wistar rats (than that in F344), the location of the microdialysis probe-membrane with respect to the area of cortical infarction differed between Wistar rats. Hence, a reliable comparison between the quantitative amount of amino acids in the dialysate from the cortical probes of both rat strains could not be made. These results, demonstrating differences in striatal excitotoxicity between Wistar and F344 rats after MCA occlusion, are the first to show interstrain differences in striatal pathophysiology of focal ischemia between these normotensive rat strains. They do however not explain why MCA occlusion results in a significantly different volume of cortical infarction between Wistar and F344 rats. The F344 strain will probably show in a more sensitive way, as compared to Wistar rats, neuroprotective effects of agents that diminish excitotoxic damage during focal cerebral ischemia.

Middle cerebral artery occlusion in Wistar and Fischer-344 rats: functional and morphological assessment of the model.

Cerebral infarction volume after occlusion of a short proximal segment of the middle cerebral artery (MCA) is reported to be different in Wistar compared to Fischer-344 (F344) rats, in both size and variability. Knowledge about the cause of these differences might enable us to explain and perhaps reduce the variation in infarct volume and create a reproducible model of focal cerebral ischemia in the rat. We investigated in Wistar and F344 rats both the effect of occlusion of a long proximal MCA segment on cerebral infarction volume, visualized by magnetic resonance imaging and histology, and the morphology of the major cerebral arteries. Occlusion of a long proximal MCA segment resulted in a striatal and a small cortical infarction in Wistar and a striatal and sizable cortical infarction in F344 rats (as is the case after occlusion of a short proximal MCA segment). In Wistar rats, however, occlusion of a long proximal MCA segment strongly reduced the variability in infarction volume in comparison to occlusion of a small proximal MCA segment. Analysis of the morphology of the major cerebral arteries showed a significantly higher number of proximal side branches of the long proximal MCA segment in Wistar rates than in F344 rats. We conclude that after short-segment proximal MCA occlusion, extreme variability in cerebral infarction volume in Wistar rats compared to F344 rats may be attributable to a significantly greater number of proximal MCA side branches in Wistar rats than in F344 rats.