Fastigial Stimulation Increases Ischemic Blood Flow and Reduces Brain Damage After Focal Ischemia

Journal of Cerebral Blood Flow and Metabolism 13:113-119 1993 The International Society of Cerebral Blood Flow and Metabolism Published by Raven Press, Ltd" New York Fastigial Stimulation Increases Ischemic Blood Flow and Reduces Brain Damage After Focal Ischemia Fangyi Zhang and Costantino Iadecola Department of Neurology, University of Minnesota Medical School, Minneapolis, Minnesota, U.S.A. Summary: Electrical stimulation of the cerebellar fastigial nucleus (FN) increases CBF and reduces brain damage after focal ischemia. We studied whether FN stimulation "protects" the brain from ischemic damage by increasing blood flow to the ischemic territory. Sprague Dawley rats were anesthetized (halothane 1-3%) and artificially ventilated through a tracheal cannula inserted transorally. CBF was monitored by a laser-doppler probe placed over the convexity at a site corresponding to the area spared from infarction by FN stimulation. Arterial pressure (AP), blood gases, and body temperature were controlled, and the electroencephalogram (EEG) was monitored. The stem of the middle cerebral artery (MCA) was occluded. After occlusion, the FN was stimulated for 6 min (1 f.la; 5 Hz; 1 s on-l s oft) while AP was maintained at 97 ± 11 mm Hg (mean ± SD) by controlled hemorrhage. Rats were then allowed to recover, and infarct volume was determined 24 h later in thionin- stained sections. In unstimulated rats (n = 7), proximal MCA occlusion reduced CBF and the amplitude of the EEG. One day later, these rats had infarcts involving neocortex and striatum. FN stimulation after MCA occlusion (n = 12) enhanced CBF and EEG recovery [61 ± 34 and 73 ± 43%, respectively at 6 min; p <.5 vs. unstimulated group; analysis of variance (ANOY A)] and reduced the volume of the cortical infarct by 48% (p <.5). In contrast, hypercapnia (Pco2 = 64 ± 4; n = 7) did not affect CBF and EEG recovery or infarct volume (p >.5). Thus, FN stimulation, unlike hypercapnia, increases CBF to the ischemic cortex, improves recovery of electrical activity, and reduces tissue damage after MCA occlusion. These findings support the hypothesis that FN stimulation reduces ischemic damage by enhancing collateral flow to the ischemic territory. Key Words: Cerebellum-Middle cerebral artery-cerebral blood flow-laser-doppler flowmetry-rat-hypercapnia. Electrical stimulation of neural pathways traversing the region of the cerebellar fastigial nucleus (FN) increases CBF globally in brain (Nakai et ai., 1983; Chid a et ai., 1989). In most regions of the cerebral cortex, the increases in CBF are not associated with corresponding increases in glucose utilization, suggesting that the vasodilation is not secondary to increased metabolic activity (Nakai et ai., 1983). Furthermore, the neocortical vasodilation is mediated by activation of central neural pathways, which, in turn, leads to the local release of the vasodilator nitric oxide (NO) (Iadecola et ai., 1983; Iadecola, 1992; Iadecola et ai., 1993). Received April 5, 1993; final revision received May 14, 1993; accepted May 21, 1993. Address correspondence and reprint requests to Dr. C. Iadecola at Department of Neurology, University of Minnesota Medical School, Box 295 UMHC, 42 Delaware Street SE, Minneapolis, MN 55455, U.S.A. Abbreviations used: AP, arterial pressure; FN, fastigial nucleus; MCA, middle cerebral artery; NO, nitrous oxide. Recent studies have demonstrated that stimulation of the FN reduces the size of the infarct resulting from middle cerebral artery (MCA) occlusion in rat (Reis et ai., 1991; Zhang and Iadecola, 1992). The "protective" effect is restricted to the cerebral cortex, the region in which the cerebrovasodilation is not associated with an increase in local metabolic demands (Zhang and Iadecola, 1992). Furthermore, the volume of tissue spared from infarction is more pronounced after distal than after proximal MCA occlusion (Zhang and Iadecola, 1992). Since the potential for collateral flow is likely to be greater after distal occlusions, these observations indicate that FN stimulation may ameliorate ischemic brain damage by enhancing collateral flow to the ischemic territory without increasing local energy requirements (Zhang and Iadecola, 1992). Therefore, we sought to determine whether FN stimulation increases CBF to ischemic tissue that is spared from infarction and, if so, whether the effect is associated with an enhanced recovery of sponta- 113

114 F. ZHANG AND C. IADECOLA neous electrical activity and a reduction in infarct volume. For comparison, we also studied whether increasing CBF by hypercapnia exerts a similar protection against cerebral ischemic damage. We found that FN stimulation, but not hypercapnia, increases CBF to the ischemic territory, enhances the recovery of the EEG, and reduces the extent of brain damage after MCA occlusion. The findings support the hypothesis that the FN protects the brain from ischemic damage by enhancing collateral perfusion and improving CBF to the ischemic territory. METHODS Methods for MCA occlusion, electrical stimulation of FN, recording and analysis of the electroencephalogram (EEG), monitoring of CBF by laser-doppler flowmetry (LDF), and determination of infarct size have been described in detail in previous publications (Iadecola, 1992; Zhang and Iadecola, 1992; 1993) and are only summarized here. General surgical procedures and procedures for FN stimulation Studies were conducted on 44 male Sprague-Dawley rats weighing 3-4 g. Anesthesia was induced with halothane (5% in 1% oxygen) administered through a facial mask. When deeply anesthetized, rats underwent transoral tracheal intubation using a method modified from that of Thet (1983). Briefly, the tongue was depressed using a spatula while the larynx was displaced posteriorly by applying slight pressure on the ventral surface of the neck. The epiglottis and vocal cords were visualized with the aid of a fiberoptic illuminator, and a polyethylene catheter (outside diameter, 2.8 mm) was gently advanced through the oral cavity into the trachea. A slight resistance was usually felt as the catheter passed through the vocal cords. After insertion, the endotracheal catheter was left in place. Both femoral arteries were then cannulated as previously described (Zhang and Iadecola, 1992). Rats were placed on a stereotaxic frame (Kopf) and artificially ventilated with 1% O2 by a mechanical ventilator (Harvard Apparatus Rodent Respirator). Ventilation with 1% O2 was necessary to compensate for any loss of oxygen-carrying capacity resulting from controlled exsanguination during FN stimulation (see Experimental protocol). Furthermore, in preliminary experiments we found that ventilation with 1% oxygen facilitates the recovery of the animals after extubation. Animals were not paralyzed. At this time, the halothane concentration was reduced to 1%. The body temperature was maintained at 37 ± O.soC by a thermostatically controlled infrared lamp connected to a rectal probe. One of the arterial catheters was used for continuous recording of arterial pressure (AP), mean AP, and heart rate, while the other catheter was used for blood sampling and controlled exsanguination during FN stimulation (see below). Small volumes of arterial blood (5 /J-l) were sampled at various times for measurement of P a2' P ac2' and ph by a blood-gas analyzer (Coming model 178). As described in detail elsewhere (Nakai et ai., 1983; Iadecola, 1992), the FN was stimulated with stereotaxically implanted microelectrodes. Stimuli were negative square-wave pulses delivered from a constant-current stimulator (Grass S88). The ground was a metal clip attached to the animal's scalp. Monitoring of CBF and EEG CBF was monitored in the cerebral cortex ipsilateral to the occluded MCA using a laser-doppler flowmeter (Vasamedic BPM 43A) (Iadecola, 1992; Iadecola et ai., 1993). A small hole, 1-2 mm in diameter, was drilled with the assistance of a dissecting scope at a site 2.5-3 mm lateral to the midline and 2-2.5 mm rostral to A. This site was found in preliminary studies to correspond to the region of the cerebral cortex that is spared from infarction by FN stimulation (see below). The dura was left intact, and the LDF probe (tip diameter,.8 mm) was positioned.5 mm above the dural surface. The analogue output of the flowmeter was fed into a DC amplifier (Grass) and displayed on the polygraph. The EEG was recorded monopolarly at a site 2 mm lateral to the midline and 1 mm rostral to the LDF probe. Techniques for quantitative analysis of the EEG have been described previously (Zhang and ladecola, 1992). Occlusion of the MCA and determination of infarct volumes Procedures for MCA occlusion and for determination of the infarct volume are identical to those published previously (Zhang and ladecola, 1992, 1993). The MCA was elevated and cauterized proximal to the origin of the lenticulostriate branches. The occlusion was extended 2-3 mm distally and the artery was transected. Twenty-four hours after MCA occlusion, rats were killed and their brains were removed. The site of CBF recording was marked, and the brain was frozen in isopentane. Coronal sections (thickness, 3 /J-m) were serially cut in a cryostat, collected at 3OO-J.l.m intervals, and stained with thionin. Infarct volume was determined using an image analyzer (MCID, Imaging Research) as previously described (Zhang and ladecola, 1992). Experimental protocol Rats were surgically prepared and the left MCA was exposed as described above. Arterial blood gases were adjusted so that arterial Pc2 was between 33 and 38 mm Hg. In animals in which the left FN was stimulated, an electrode was inserted into the cerebellum at a site 5 mm rostral to the obex and.8 mm lateral to the midline (Nakai et ai., 1983). The most active pressor site in FN was then localized using procedures described in detail elsewhere (Iadecola, 1992). Once the most active site in FN was localized, the electrode was left in place and procedures for occlusion of the MCA were begun. Occurrence of neocortical ischemia was indicated by a drop in CBF and a 6-8% reduction in the amplitude of the EEG (Fig. 1). In animals in which the FN was stimulated, stimulation was begun 3-5 min after MCA occlusion. The FN was stimulated with intermittent trains of stimuli (1 s on-l s off; 5 Hz; 75-1 J.l.A), and the evoked elevations in AP were offset by the slow removal of small amounts (1-2 ml) of arterial blood. Because of this procedure, AP did not differ between stimulated and unstimulated rats (Table 1). The FN was stimulated for 1 h. In animals in which the effect of hypercapnia was studied, 5% CO2 was added to the circuit of the respirator 3-5 min after MCA occlusion. Hypercapnia was maintained for 1 h. At the end of the period of FN stimulation or hypercapnia, catheters were removed and wounds infiltrated with lidocaine and su- J Cereb Blood Flow Metab, Vol. 13, No.6, 1993

NEUROGENIC VASODILATION AND FOCAL ISCHEMIA 115 (A) FN stirn RESULTS (8) CBF 1 % of Cont. EEG 1 flv I I \'--- occlusion + 1 sec MCA occlusion 1 Min 6 Min FIG. 1. Experimental protocol for occlusion of the middle cerebral artery (MCA) with monitoring of cerebral blood flow (CBF) and electroencephalogram (EEG). A: The MCA was elevated, cauterized, and transected proximal to the origin of the lenticulostriate branches (MCA occlusion). CBF was monitored by using a laser-doppler probe. The position of the probe was chosen in preliminary studies to correspond to the posterior border of the infarcted territory. The EEG was monitored using a screw electrode inserted 1 mm rostral to the CBF probe. At the end of the experiment, the position of the CBF probe was marked and animals were allowed to recover. Twenty-four hours later, infarct volume was determined on thionin-stained sections using an image analysis system. The shaded area outlined by the dashed line illustrates the distribution of the infarcted tissue 24 h after MCA occlusion. Note that the CBF probe was located over tissue that underwent infarction. In some animals, the cerebellar fastigial nucleus (FN) was stimulated through microelectrodes positioned stereotaxically (FN stim). 8: Effect of MCA occlusion on CBF and EEG in the ischemic hemisphere. Occlusion of the MCA produced a drop in CBF and in the amplitude of the EEG. The depression in CBF and EEG was sustained with only a slight recovery over the next 6 min. Using this model, the time course of the changes in CBF and EEG occurring in the critical phases after MCA occlusion can be correlated with the histological outcome of the tissue in which CBF and EEG were recorded. tured. Animals were extubated and carefully observed during the postoperative period for the occurrence of seizures. Twenty-four hours later, rats were killed for determination of infarct volume. Data analysis Data are expressed as means ± SD. Comparisons among multiple groups were statistically evaluated by analysis of variance and Tukey's test (Systat). Differences were considered significant for p <.5. Effects of FN stimulation or hypercapnia on CBF in sham-occluded rats In unstimulated rats in which the MCA was exposed but not occluded (sham occlusion), CBF was stable throughout the period of monitoring (Fig. 2). With FN stimulation (1!-LA; 5 Hz), CBF increased steadily and remained elevated throughout the stimulation period ( + 13 ± 38% at 1 min; Fig. 2). Similarly, hypercapnia (P aco2 = 65 ± 4 mm Hg; n = 6) resulted in a sustained increase in CBF that was comparable in magnitude to that elicited by FN stimulation (p <.5 vs. control; p >.5 vs. FN stimulation). In these sham-occluded groups, a small area of necrosis in the cortex adjacent to the surgical site was observed 24 h later (Table 2). Effect of FN stimulation or hypercapnia on CBF and EEG after MCA occlusion In unstimulated rats (n = 7), proximal occlusion of the MCA resulted in an immediate drop in CBF ( -86 ± 17%) and a reduction in the amplitude of the EEG (-67 ± 1%; Fig. 1). Sixty minutes later, CBF and EEG had recovered by 11 ± 5 and 25 ± 27%, respectively (Fig. 3). Twenty-four hours later, these animals had large infarcts involving neocortex and striatum (Table 2). In the group of rats in which the FN was stimulated (n = 12), MCA occlusion resulted in drops in CBF and EEG amplitude similar to those in the unstimulated group (p >.5). However, the recovery of CBF and EEG was significantly greater than in the unstimulated group (p <.5 at 4 and 6 min; Fig. 3). In these rats, the size of the neocortical infarct 24 h later was 48 ± 36% smaller than in the unstimulated group (p <.5; Table 2). In contrast to FN stimulation, hypercapnia (P aco2 = 64 ± 4 mm Hg) failed to influence the recovery of CBF and EEG (Fig. 3) or to reduce the size of the infarct resulting from MCA occlusion (Table 2). Relationship between CBF probe and tissue spared from infarction In all rats, the position of the CBF probe was marked so that location of the probe relative to the area of stroke could be determined histologically. The distribution of the infarcted tissue in reference to the location of the CBF probe in representative cases is illustrated in Fig. 4. In all stimulated rats, the tissue over which the probe was positioned did not undergo infarction (Fig. 4). In contrast, in unstimulated animals and in animals subjected to hypercapnia, the tissue located under the probe did undergo infarction (Fig. 4). Therefore, the pattern of CBF recovery observed with FN stimulation re- J Cereb Blood Flow Metab, Vol. 13, No.6, 1993

116 F. ZHANG AND C. IADECOLA TABLE 1. Body weight, arterial pressure, and blood gases in rats with or without occlusion of the middle cerebral artery Sham-operated MCA-occluded UN FNS He UN FNS He Arterial pressure (mm Hg) 95 ± 4 97 ± 11 95 ± 12 94 ± 7 97 ± 11 97 ± 11 Pcoz (mm Hg) 35 ± 2 36 ± 1 65 ± 4a 35 ± 2 35 ± 2 64 ± 4a POz (mm Hg) 452 ± 67 427 ± 5 512 ± 52 469 ± 15 467 ± 74 486 ± 42 ph 7.44 ±.7 7.46 ±.3 7.25 ± O.3a 7.44 ±.5 7.48 ±.3 7.27 ± O.4a Body weight (g) 355 ± 58 344 ± 22 341 ± 27 314 ± 41 322 ± 32 313 ± 16 No. 6 6 6 7 12 7 MCA, middle cerebral artery; UN, unstimulated; FNS, FN stimulated; He, hypercapnia. Values are means ± SD. a p <.1 vs. unstimulated and FN-stimulated groups (analysis of variance and Tukey's test). flects that of the ischemic region that was spared from infarction. DISCUSSION We sought to determine whether the reduction in cerebral ischemic damage afforded by FN stimulation results from an increase in CBF to the ischemic tissue. To achieve this goal, we developed an experimental protocol for occlusion of the MCA while monitoring CBF in the ischemic hemisphere under conditions of controlled AP, blood gases, and body temperature. Using this method, we were able to correlate, in each animal, the changes in CBF during the critical period after MCA occlusion with the histological outcome of the tissue in which CBF was monitored. We found that FN stimulation enhanced the recovery of CBF in the ischemic tissue that did not undergo infarction. In contrast, hyper- " 'E o u - o u.. III U 3 25 2 15 1 5 // /1 / / //.. / // ---- Control (n=6). FN Stimulation (n=6) - -.. - Hypercapnia (n=6) p<.5 t t,f.,/ / : " - 1 --------1. ::::.. t OL------- o 2 4 6 Time (min) FIG. 2. Effect of FN stimulation or hypercapnia on CBF in rats in which the middle cerebral artery was exposed but not occluded (sham occlusion). In unstimulated rats, CBF tended to increase slightly over the 6 min of monitoring. However, these CBF changes were not statistically significant (p >.5, analysis of variance). FN stimulation (1 f.la; 5 Hz; 1 s on-1 s off) or hypercapnia (Pco4 = 65 ± 4) produced sustained increases in CBF (p <.5 vs. normocapnic unstimulated group) that were similar in magnitude. capnia, a vasodilator stimulus that produces increases in CBF comparable with those accompanying FN stimulation, failed to increase CBF in the ischemic territory or to reduce infarct size. The results indicate that FN stimulation, unlike hypercapnia, can improve ischemic blood flow and limit the extent of the brain damage resulting from focal ischemia. The effect of FN stimulation on the size of the infarct and on CBF in the ischemic territory is not due to differences in AP, blood gases, or body temperature between unstimulated and stimulated animals; these parameters were carefully controlled and did not differ between groups. Although brain temperature was not measured in this study, it is unlikely that the protective effect of FN stimulation is due to brain hypothermia (Morikawa et al., 1992a); FN stimulation increases CBF and, consequently, should increase brain temperature, and an increase in brain temperature does not ameliorate but worsens focal ischemic damage (Morikawa et ai., 1992a). Similarly, the protective effect of FN stimulation is unlikely to result from changes in plasma glucose, because FN stimulation leads to hyperglycemia (Nakai et ai., 1983), a condition that worsens focal ischemic damage (Prado et ai., 1988). An important question concerns the mechanisms by which FN stimulation ameliorates focal ischemic damage. In analogy with other interventions for brain protection (Choi, 199), FN stimulation could reduce tissue damage by improving CBF to the ischemic tissue and decreasing the degree of oligemia (hemodynamic mechanism). Alternatively, FN stimulation might increase the resistance of brain cells to ischemic damage by releasing protective agents in the ischemic territory, possibly neurotransmitters (cytoprotective mechanism) (Reis et al., 1991). The finding that FN stimulation increases CBF in the ischemic territory strongly suggests that the mechanism of the protection is "hemodynamic." However, the vascular mechanisms by which FN stimulation increases CBF to the isch- J Cereb Blood Flow Metab. Vol. 13, No.6, 1993

NEUROGENIC VASODILATION AND FOCAL ISCHEMIA 117 TABLE 2. Effect of FN stimulation or hypercapnia on the volume of the infarct resulting fr om occlusion of the MeA in anesthetized rat Sham-operated MeA-occluded UN FNS He UN FNS He Total 18 ± 4 12 ± 5 15 ± 5 39 ± 31 183 ± 82a 33 ± 45 Neocortex 18 ± 4 12 ± 5 15 ± 5 224 ± 32 116 ± 8a 217 ± 5 Striatum 58 ± 1 59 ± 7 65 ± 9 No. 6 6 6 7 12 7 Values are means ± SD (mm3). a p <.1 vs. MeA-occluded unstimulated and hypercapnia groups (analysis of variance and Tukey's test). ernie territory remain to be elucidated. Since FN stimulation increases CBF globally, it is conceivable that the vasodilation enhances collateral flow to the areas surrounding the ischemic core. Thus, the protective effect may depend on blood flow from cortical anastomoses between distal MCA branches and adjacent vessels not involved by ischemia (Coyle and Jokelainen, 1982). Another possibility is that FN stimulation does not permanently protect the tissue from ischemic damage but only delays the occurrence of infarction. Consequently, 24 h after MCA occlusion, the infarcted area would appear smaller in stimulated than in unstimulated rats. This possibility cannot be excluded on the basis of our study; however, if it were the case, the effects of FN stimulation would have to be long lasting and, as such, difficult to explain on the basis of stimulation-evoked, transient hemodynamic changes. Which mediators are responsible for the increases of CBF in the ischemic territory? Recent data indicate that NO participates in the neocortical cerebrovasodilation elicited by FN stimulation (Iadecola, 1992; Iadecola et al., 1993). It is therefore conceivable that local release of NO, a potent vasodilator and inhibitor of platelet aggregation (Moncada, 1992), improves blood flow in the ischemic territory. Indeed, there is a growing body of evidence suggesting that NO is beneficial in the early stage of focal ischemia. For example, intracarotid administration of NO donors increases ischemic blood flow and reduces the size of the stroke resulting from MCA occlusion (Zhang and Iadecola, 1993). In addition, administration of the NO precursor L-arginine attenuates the infarct resulting from MCA occlusion (Morikawa et al., 1992b). Therefore, FN stimulation may increase CBF in the ischemic territory by increasing NO production. On the other hand, NO could protect the tissue from infarction by mechanisms independent of its ability to produce vasodilation and inhibit platelet aggregation. NO has been shown to block N-methyl-oaspartate (NMDA) receptors in vitro, possibly by 12 1 e- 8 >- > II: 6 LL. m 4 (,) >- > II: <.!' w w 2 12 1 8 6 4 2 - -Q- --Ir- --.. Hypercapnia _-- p<o.5 Control (n=7) FN Stimulation (n=12) (n=7) I ---------- 2 4 6 1 I - --1, ------------------ -i / 1! I, 2 4 6 Time (min) FIG. 3. Effect of FN stimulation or hypercapnia on the recovery of cerebral blood flow (A) and EEG (8) after occlusion of the middle cerebral artery in anesthetized rats. Stimulation of the FN, but not hypercapnia, enhanced the recovery of EEG and CBF after MCA occlusion (p <.5, analysis of variance). The mean reduction in CBF, averaged over 6 min, was -88 ± 11% in unstimulated rats, -85 ± 13% in hypercapnic rats, and -53 ± 31 % in FN-stimulated rats (p <.1 vs. unstimulated and hypercapnic rats). (A) (8) J Cereb Blood Flow Metab, Vol. ]3, No.6, 1993

118 F. ZHANG AND C. IADECOLA CBF '--' Imm FIG. 4. Stimulation of the fastigial nucleus (middle) but not hypercapnia (right), led to smaller infarcts after occlusion of the middle cerebral artery (MCA). The position of the laser Doppler probe used to monitor cerebral blood flow (CBF) after MCA occlusion is illustrated. In unstimulated and in hypercapnic rats, the CBF probe overlies infarcted tissue; in rats subjected to FN stimulation, the probe is located over tissue that did not undergo infarction. The unstimulated subject is shown at the left. acting on the redox modulatory site (Lei et ai., 1992). Therefore, NO, like NMDA receptor antagonists, might protect neurons from anoxic-ischemic damage (Choi, 199). Whether NO attenuates NMDA-mediated neuronal injury, however, remains highly controversial; other studies have indicated that NO promotes excitotoxic neuronal damage (see Dawson et ai., 1991). Thus, further studies will be needed to clarify the role of NO in anoxicischemic damage. In apparent contrast with the hypothesis that NO is responsible for the protective effect of FN stimulation is the observation that hypercapnia, a vasodilator response that may also depend on NO production (for review, see Iadecola, 1993), does not increase CBF in the ischemic territory and does not reduce focal ischemic damage in rat, as found in the present study and by Jones et ai. (1989). However, there are important differences between the vasodilation elicited by hypercapnia and FN stimulation that should be emphasized. Although both responses are attenuated by NO synthase inhibitors (Iadecola et ai., 1993), the source of NO is different in these two conditions. During FN stimulation, NO is probably generated by cortical neurons containing NO synthase (Iadecola, 1992, 1993), while in hypercapnia the source of NO is likely to be in nonneuronal cells (Iadecola et ai., 1987; N orins et ai., 1992). Therefore, the difference between the effect of hypercapnia and the effect of FN stimulation on ischemic damage suggests that the site of production may be an important factor for the effect of NO in cerebral ischemia. On the other hand, the discrepancy may indicate that NO is not responsible for the protection exerted by the FN and that other factors may be involved. In conclusion, we have demonstrated that FN stimulation increases CBF to the region that is spared from focal infarction. While the mechanisms of the effect remain to be fully elucidated, the findings strongly suggest that FN stimulation protects the brain from focal ischemia by increasing local production of the potent vasodilator NO and enhancing collateral flow to the ischemic territory. Thus, intrinsic neural pathways acting through NO modulate the expression of cerebral ischemic damage. Acknowledgment: Supported by Grants-in-Aid from the American Heart Association (Minnesota) and by the Dr. Louis Sklarow Memorial Fund. F.Z. is a Fellow of the American Heart Association (Minnesota). C.1. is an Established Investigator of the American Heart Association (National). REFERENCES Chida K, Iadecola C, Reis OJ (1989) Global reduction in cerebral blood flow and metabolism elicited from intrinsic neurons of fastigial nucleus. Brain Res 5:177-192 Choi OW (199) Cerebral hypoxia: Some new approaches and unanswered questions. J Neurosci 1:2493-251 Coyle P, Jokelainen PT (1982) Dorsal cerebral arterial collaterals of the rat. Anat Rec 23:397-44 Dawson VL, Dawson TM, London ED, Bredt OS, Snyder SH (1991) Nitric oxide mediates glutamate neurotoxicity in primary cortical cultures. Proc Natl Acad Sci USA 88:6368-6371 Iadecola C (1992) Nitric oxide participates in the cerebrovasodilation elicited from cerebellar fastigial nucleus. Am J Physiol 263:RI156-RI161 Iadecola C (1993) Regulation of the cerebral microcirculation during neural activity: Is nitric oxide the missing link? Trends Neurosci 16:26-214 Iadecola C, Mraovitch S, Meeley M, Reis (1983) Lesions of the basal forebrain in rat selectively impair the cortical vasodilation elicited from cerebellar fastigial nucleus. Brain Res 279:41-52 Iadecola C, Arneric S, Baker H, Tucker L, Reis (1987) Role of local neurons in the cerebrocortical vasodilation elicited from cerebellum. Am J Physiol 252:R182-RI91 Iadecola C, Zhang F, Xu X (1993) Role of nitric oxide synthasecontaining vascular nerves in cerebrovasodilation elicited from cerebellum. Am J Physiol 264:R738-R746 Jones SC, Bose B, Furlan AJ, Friel HT, Easley KA, Meredith MP, Little JR (1989) CO2 reactivity and heterogeneity of cerebral blood flow in ischemic, border zone, and normal cortex. Am J Physiol 26:H473-H482 Lei SZ, Pan ZH, Aggarwal SK, Chen HS, Hartman J, Sucher NJ, Lipton SA (1992) Effect of nitric oxide production on the redox modulatory site of the NMDA receptor-channel complex. Neuron 8: 187-199 J Cereb Blood Flow Metab. Vol. 13, No.6, 1993

NEUROGENIC VASODILATION AND FOCAL ISCHEMIA 119 Moncada S (1992) The 1992 Vlf von Euler Lecture. The L-arginine: nitric oxide pathway. Acta Physiol Scand 145:21-227 Morikawa E, Ginsberg MD, Dietrich WD, Duncan RC, Kraydieh S, Globus MY, Busto R (1992a) The significance of brain temperature in focal cerebral ischemia: histopathological consequences of middle cerebral artery occlusion in the rat. J Cereb Blood Flow Metab 12:38--389 Morikawa E, Huang A, Moskowitz M (1992b) L-Arginine decreases infarct size caused by middle cerebral arterial occlusion in SHR. Am J Physiol 263:HI632-HI635 Nakai M, Iadecola C, Ruggiero DA, Tucker LW, Reis DJ (1983) Electrical stimulation of cerebellar fastigial nucleus increases cerebrocortical blood flow without change in local metabolism: Evidence for an intrinsic system in brain for primary vasodilation. Brain Res 26:35-49 Norins NA, Wendelberger K, Hoffman RG, Keller PA, Madden JA (1992) Effects of indomethacin on myogenic contractile activation and responses to changes in O2 and CO2 in isolated feline cerebral arteries. J Cereb Blood Flow Metab 12:866--872 Prado R, Ginsberg MD, Dietrich WD, Watson BD, Busto R, (1988) Hyperglycemia increases infarct size in collaterally perfused but not end-arterial vascular territories. J Cereb Blood Flow Metab 8:186-192 Reis OJ, Berger SB, Underwood MD, Khayata M (1991) Electrical stimulation of cerebellar fastigial nucleus reduces ischemic infarction elicited by middle cerebral artery occlusion in rat. J Cereb Blood Flow Metab 11:81--818 Thet LA (1983) A simple method of intubating rats under direct vision. Lab Anim Sci 33:368-369 Zhang F, Iadecola C (1992) Stimulation of the fastigial nucleus enhances EEG recovery and reduces tissue damage after focal cerebral ischemia. J Cereb Blood Flow Metab 12:962-97 Zhang F, Iadecola C (1993) Nitroprusside improves blood flow and reduces brain damage after focal ischemia. Neuroreport 4:559-562 J Cereb Blood Flow Metab, Vol. 13, No.6, 1993