Moderate Hypothermia Reduces Hypotensive,But Not Hypercapnic Vasodilation of Pial Arterioles in Rats

MATERIALS AND METHODS

Male Sprague-Dawley rats (350 to 400 g) were anesthetized by an intraperitoneal injection of α-chloralose (50 mg/kg) and urethane (750 mg/kg), both of which were supplemented as necessary during the procedure. The femoral arteries on both sides were cannulated to monitor blood pressure and to withdraw blood to induce hypotension. The rats were given a tracheostomy and mechanically ventilated. End-tidal CO₂ was continuously monitored using a CO₂ analyzer (Respina IH31, San-ei, Tokyo, Japan) and maintained at appropriate levels. Arterial blood pH and gases were measured with a pH/blood gas analyzer (168 pH blood gas system, Coming, Medfield, MA, U.S.A.). Rectal temperature was maintained at 37°C.

A closed cranial window was constructed as described previously (Morii et al., 1986) over the right parietal cortex. Pial microvessels were then observed in situ using a microscope (Nikon, Tokyo, Japan) fitted with a video camera system (Series CCD C2400 system, Hamamatsu Photonics, Hamamatsu, Japan). Briefly, the head of the rat was fixed in a frame and three polyethylene tubes were embedded in a ring of dental acrylic applied to the skull to suffuse the window with artificial CSF and to monitor intracranial pressure. A thermosensor was also embedded in the acrylic to monitor temperature in the window. Craniectomy (5 × 4 mm) was then performed over the right parietal cortex (1 to 6 mm posterior to the bregma and 1 to 5 mm lateral to the midline) in the center of the acrylic, and the dura—arachnoid complex was incised while the pial surface was superfused with artificial CSF. A glass coverslip was placed on the acrylic ring and sealed with cyanoacrylate. The pial arteriolar diameter was continuously measured using a video dimension analyzer (Model C1170, Hamamatsu Photonics).

RESULTS

Physiologic parameters and CO₂ reactivities (dilation [%]/Paco₂ increase [mm Hg]) are shown in Table 1. The Paco₂ is indicated at corrected (measured at the animals' actual rectal temperature) and uncorrected temperatures (measured at 37°C). The pial arterioles of the alpha-stat group constricted approximately 11 % as the temperature was lowered, whereas the baseline diameter of pial arterioles in the pH-stat group did not change. Carbon dioxide reactivity was well preserved in both hypothermic groups. In contrast, vasodilation during induced hypotension was significantly reduced in the hypothermic groups (P < 0.01). In the pH-stat group, the arterioles did not show any significant changes until the MAP was decreased to 60 mm Hg. The reduction in hypotensive vasodilation was larger in the pH-stat than the alpha-stat group (P < 0.01, Fig. 1).

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FIG. 1.

Changes in the pial arteriolar diameter were plotted as a function of changes in mean arterial pressure. Vasodilation associated with induced hypotension in hypothermic groups was reduced compared with that of the normothermic group (P < 0.01). The degree of attenuation in hypotensive vasodilation was greater under conditions of pH-stat than alpha-stat management (P < 0.01). Data represent the mean ± SD. *P < 0.05 compared with baseline diameter by one-way analysis of variance followed by Bonferroni/Dunn test.

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TABLE 1.

Physiologic parameters and CO₂ reactivity

	Control (n = 7)	alpha-stat (n = 8)		pH-stat (n = 7)
		Before hypothermia	Hypothermia	Before hypothermia	Hypothermia
Rectal temperature (°C)	37.0 ± 0.4	37.3 ± 0.3	30.0 ± 0.1	37.3 ± 0.4	30.0 ± 0.4
Window temperature (°C)	36.4 ± 0.2	36.4 ± 0.4	30.3 ± 0.4	36.6 ± 0.3	30.2 ± 0.4
pH
Temperature corrected	7.43 ± 0.03	7.44 ± 0.03	7.55 ± 0.05	7.45 ± 0.04	7.40 ± 0.05
Temperature uncorrected	ND	ND	7.44 ± 0.05	ND	7.30 ± 0.05
Pao2 (mm Hg)	138 ± 16	144 ± 16	154 ± 18	139 ± 30	129 ± 27
Paco2 (mm Hg)
Temperature corrected	36 ± 2	35 ± 1	24 ± 1	34 ± 3	35 ± 4
Temperature uncorrected	ND	ND	33 ± 2	ND	49 ± 6
Mean arterial pressure (mm Hg)	121 ± 10	119 ± 10	124 ± 12	129 ± 4	131 ± 13
Baseline diameter (µm)	44 ± 7	46 ± 7	41 ± 8	42 ± 11	43 ± 11
CO2 reactivity (%/mm Hg)	1.2 ± 0.2	1.1 ± 0.3	1.1 ± 0.4	1.1 ± 0.2	1.0 ± 0.3

Data represet the mean ± SD. ND, not determined.

DISCUSSION

Autoregulation is the intrinsic ability to maintain constant blood flow despite changes in blood pressure. At the level of individual vessels, cerebral blood flow is autoregulated by arteriolar dilation in response to a fall in perfusion pressure. We examined autoregulation under moderate hypothermia using a closed cranial window that allowed direct observation of pial arterioles in situ under temperature-controlled conditions. We found that pial arteriolar dilation in the face of hypotension is significantly reduced under hypothermia.

The acid—base balance during hypothermia was maintained by alpha-stat and pH-stat management as described previously (Swain, 1988). There has been considerable controversy over which procedure is physiologically more suitable for nonhibernating homeotherms and more effectively prevents cerebral damage after cardiac surgery (Stephan et al., 1992; Kern and Greeley, 1995; Burrows, 1995). In the present study, although ventilation was decreased during cooling and temperature-uncorrected Paco₂ increased to 50 mm Hg (see Table 1) in the pH-stat group, the baseline diameter of the arterioles did not change. In contrast, the baseline diameter in the alpha-stat group decreased although temperature-uncorrected Paco₂ was stable. Changes in the baseline diameter during the introduction of hypothermia was compatible with temperature-corrected Paco₂ (decreased in alpha-stat and constant in pH-stat management). These findings indicate that the pH-stat condition is relatively hypercapnic compared with the alpha-stat condition, but not hypercapnic as observed when the Paco₂ was 50 mm Hg. Thus, temperature-corrected blood gas analysis may be relevant to cerebrovascular physiology under hypothermia.

Pial arteriolar dilation in the face of hypotension was reduced under hypothermia although hypercapnic vasodilation was preserved in this study. Thus, we believe that impairment was specifically induced by faulty autoregulation, and not by nonspecific decline of cerebrovascular reactivity. In addition, the degree of impaired autoregulation was greater in the pH-stat, than in the alpha-stat, condition, These results were consistent with the findings from a study of the effects of moderate hypothermia and acid-base management on cerebral autoregulation under nonbypass conditions, using laser—Doppler flowmetry (Verhaegen et al., 1993).

However, when interpreting these results, the following points should be considered, First, hypotension was induced by withdrawing blood, which is associated with increases in vasoactive substances such as catecholamines, which may in turn affect pial arteriolar reactivity, Whether there is any difference in release of vasoactive compounds during induced hypotension between the normothermic and hypothermic conditions is unknown. Second, hypotensive vasodilation may have shifted toward the lower absolute levels of MAP in the hypothermic groups, However, it is unlikely that further dilation (maximal dilation) occurred below an MAP of 20 mm Hg in the hypothermic groups, which is beyond the MAP levels we examined, and thus we believe that hypotensive vasodilation was reduced, not simply shifted, in the hypothermic groups.

There are several possible mechanisms involved in altered autoregulation. The attenuation of cerebral metabolism induced by hypothermia, resulting in reduced tissue oxygen demands, may result in reduced hypotensive vasodilation, An oxygen-sensitive mechanism for cerebral blood flow autoregulation has been proposed in which tissue hypoxia would be responsible for arteriolar dilation through the liberation of vasodilator substances (Kontos and Wei, 1985). The stimulus for vasodilation (tissue hypoxia) may be relatively blunt in the hypothermic brain in which cerebral metabolic rates for oxygen are markedly decreased (approximately 65% of normal at 30°C according to Hägerdal et al., 1975), resulting in reduced production of mediators for vasodilation.

Another possible mechanism involves an increase in blood viscosity induced by hypothermia (Chen and Chien, 1978). Change in blood viscosity is an important factor for regulation of cerebral blood flow (Muizelaar et al., 1986) by changes in shear stress to the endothelium, with accompanying changes in release of endothelium-derived relaxing factor. This may contribute to alteration in cerebral blood flow autoregulation.

The involvement of other potential mediators of altered autoregulation, including adenosine (Berne et al., 1974; Kontos and Wei, 1985), calcitonin gene-related peptide (Lee et al., 1993), parasympathetic nerves (Koketsu et al., 1992), and nitric oxide (Tanaka et al., 1993), should also be considered. However, these mechanisms may not completely explain why autoregulation is more profoundly altered under pH-stat conditions as shown by this study and those of others (Murkin et al., 1987; Verhaegen et al., 1993). When considering this issue, two possibilities need to be taken into account, if the oxygen-dependent mechanism is assumed to play a major role in hypotensive vasodilation.

First, the baseline diameter changes seen in the present study (i.e., the pial arterioles constricted in the alpha-stat group whereas the baseline diameter did not change in the pH-stat group) suggest that the pH-stat condition is one of relative hyperemia for the reduced cerebral metabolism during hypothermia compared with the alpha-stat condition. We speculate that the increase in tissue oxygen demands in response to induced hypotension are relatively small in a brain with relative hyperemia under pH-stat conditions. Second, should the CMRO₂ be lower in the pH-stat condition than in the alpha-stat condition? The finding that Paco₂ hypercarbia of 55 to 68 mm Hg reduces CMRO₂ 25% to 50% during hypothermia (26° to 28°C) is in line with this concept (Prough et al., 1990). In contrast, other studies have demonstrated no difference in CMRO₂ between the two types of acid—base management at 26° to 27°C (Murkin et al., 1987; Stephan et al., 1992; Hindman et al., 1993). The role of this mechanism remains to be clarified. Another important consideration regarding this issue is that pH-stat management may cause relative hypercapnia and vasodilation, which limits further dilation in response to hypotension (Murkin et al., 1987; Verhaegen et al., 1993). However, our findings conflict with this concept because the baseline diameter of pial arterioles did not change, and the hypercapnic response was preserved after pH-stat conditions were introduced, indicating that the vessels were not yet dilated.

We demonstrated that moderate hypothermia and acid—base management alters the autoregulatory response, although the mechanism is not yet fully understood. The contribution of altered cerebrovascular reactivity to neuroprotection is currently under investigation.