Sage Journals: Discover world-class research

Abstract

Alterations in cerebral autoregulation and cerebrovascular reactivity after traumatic brain injury (TBI) may increase the susceptibility of the brain to secondary insults, including arterial hypotension. The purpose of this study was to evaluate the consequences of mild hemorrhagic hypotension on hemodynamic and histopathologic outcome after TBI. Intubated, anesthetized male rats were subjected to moderate (1.94 to 2.18 atm) parasagittal fluid–percussion (FP) brain injury. After TBI, animals were exposed to either normotension (group 1: TBI alone, n = 6) or hypotension (group 2: TBI + hypotension, n = 6). Moderate hypotension (60 mm Hg/30 min) was induced 5 minutes after TBI or sham procedures by hemorrhage. Sham-operated controls (group 3, n = 7) underwent an induced hypotensive period, whereas normotensive controls (group 4, n = 4) did not. For measuring regional cerebral blood flow (rCBF), radiolabeled microspheres were injected before, 20 minutes after, and 60 minutes after TBI (n = 23). For quantitative histopathologic evaluation, separate groups of animals were perfusion-fixed 3 days after TBI (n = 22). At 20 minutes after TBI, rCBF was bilaterally reduced by 57% ± 6% and 48% ± 11% in cortical and subcortical brain regions, respectively, under normotensive conditions. Compared with normotensive TBI rats, hemodynamic depression was significantly greater with induced hypotension in the histopathologically vulnerable (P1) posterior parietal cortex (P < 0.01). Secondary hypotension also increased contusion area at specific bregma levels compared with normotensive TBI rats (P < 0.05), as well as overall contusion volume (0.96 ± 0.46 mm³ vs. 2.02 ± 0.51 mm³, mean ± SD, P < 0.05). These findings demonstrate that mild hemorrhagic hypotension after FP injury worsens local histopathologic outcome, possibly through vascular mechanisms.

Keywords

Autoregulation Fluid-percussion injury Hypoxia Pathology Rat Regional cerebral blood flow

Posttraumatic secondary events—including ischemia, hypoxia, hypotension, hyperthermia, and intracranial hypertension—are commonly observed in head-injured patients (Bouma and Muizelaar, 1992; Chesnut et al., 1993; Golding et al., 1999; Kelly et al., 1997; Marion et al., 1991; Muizelaar et al., 1984; Obrist et al., 1984). It is also well documented that secondary insults contribute to the mortality and morbidity associated with clinical (Chesnut et al., 1993; Muizelaar et al., 1984) and experimental (Bramlett et al., 1999; Cherian et al., 1996; Clark et al., 1997; Davis et al., 1998; DeWitt et al., 1992a, 1992b, 1995; Dietrich et al., 1996;Golding et al., 1999; Ishige et al., 1987; Jenkins et al., 1989; Kroppenstedt et al., 1999; Nawashiro et al., 1995; Schmoker et al., 1992) traumatic brain injury (TBI). Cerebrovascular perturbations, including structural (Cortez et al., 1989; Dietrich et al., 1994; Povlishock et al., 1978; Wei et al., 1980) and hemodynamic perturbations (Armstead and Kurth, 1994; Davis et al., 1998; DeWitt et al., 1986, 1992a, 1992b; Dietrich et al., 1996a, 1998; Yamakami and McIntosh, 1991), are a common consequence of experimental TBI and may underlie the increased sensitivity of the posttraumatic brain to secondary insults.

Alterations in global and regional cerebral blood flow (rCBF) have been documented in patients with head injuries (Bouma and Muizelaar, 1992; Marion et al., 1991; Martin et al., 1997; Obrist et al., 1984). In models of moderate fluid–percussion brain injury (FP), rCBF reductions are common in the mild to moderate range (Yamakami and McIntosh, 1991; DeWitt et al., 1986, 1992a, 1992b; Dietrich et al., 1996a), whereas severe FP injury leads to ischemic levels of rCBF (Dietrich et al., 1998). In contrast, models of controlled cortical impact injury are known to produce regional cerebral ischemia (Bryan et al., 1995; Dietrich et al., 1998; Kochanek et al., 1995; Nilsson et al., 1996). In human TBI, histopathologic damage in selectively vulnerable brain regions, including the hippocampus, is observed (Adams et al., 1982; Graham et al., 1978). Thus, under specific experimental and clinical conditions, posttraumatic ischemia is an acute response to TBI and may participate in the pathophysiology of this injury.

Dysfunction of cerebral autoregulation also has been reported in animal models of TBI (Golding et al., 1999). In the acute stages of injury, several studies have reported that TBI impairs and/or abolishes the CBF response to induced hypotension, increased CO₂, or other vasoactive substances (Armstead, 1998; Engelborghse et al., 2000; Law et al., 1996; Lewelt et al., 1980; Newell et al., 1996; Wei et al., 1980). DeWitt and colleagues (1992b) demonstrated that global and regional autoregulation are absent in response to hemorrhagic hypotension after midline FP brain injury in cats. Recently, Giri and colleagues (2000) reported that after cortical impact injury in rats, rCBF also is further reduced at the impact site when the injured brain is confronted with a secondary ischemic insult. This impairment in rCBF maintenance with respect to changes in Pco₂ and cerebral perfusion pressure also have been observed in the traumatically brain-injured patient (Muizelaar et al., 1984; Newell et al., 1996).

There are two prevailing mechanisms that have been proposed in mediating autoregulation. One is a metabolic component whereby neuronal activity induces the release of some neurochemicals (for example, nitric oxide and adenosine) that vasodilate arterioles (Golding et al., 1999). Acute disturbances in glucose metabolism or strong vasoconstrictors such as endothelin have been reported in experimental models of TBI and might participate in these abnormalities (Ginsberg et al., 1997; Yoshino et al., 1991). Consequently, experimental TBI models may have a propensity for autoregulatory dysfunction by depleting a vasodilator or eliciting an abnormal increase in a vasoconstrictor, resulting in the loss of vascular coupling.

The other proposed autoregulatory mechanism is myogenic. This cellular response is because of the interaction of smooth muscle cells to changes in vascular wall tension. Thus, contraction of the vessel wall will occur if perfusion pressure increases and the opposite occurs if pressure decreases. Golding and colleagues (1998) have demonstrated temporal changes in the myogenic response in the middle cerebral artery (MCA) after severe cortical impact injury. They found that although the mechanisms of contraction were present, changes existed in the interaction of myogenic tone or response indices to pressure at 24 hours after trauma. Recently, Mathew and colleagues (1999) reported that cerebral arterial vasodilatory responses to progressive reductions in intraluminal pressure also are affected after moderate central FP injury. In Mathew's study, MCA segments harvested from traumatized rats exhibited abnormal vasodilation during pressure reductions. When regional autoregulation is dysfunctional, secondary insults, such as hypotension, might lead to regional ischemia and worsen outcome (Bouma and Muizelaar, 1992).

Posttraumatic hypoxia has been reported to worsen outcome after TBI (Bramlett et al., 1999; Clark et al., 1997; Ishige et al., 1987; Nawashiro et al., 1995). In one study, 30-minute hypoxia after moderate FP brain injury significantly increased cortical contusion volume and the frequency of CA1 hippocampal neuronal damage compared with normoxic TBI rats (Bramlett et al., 1999). Because TBI hypoxic animals in that study also exhibited mild hypotension, the interpretation of which specific mechanisms were responsible for increased damage (that is, hypoxia vs. hypotension) was unclear. Hypotensive episodes occur in 15% to 35% of patients with severe head injuries and may increase mortality and morbidity associated with that injury (Chesnut et al., 1993). Therefore, the current study was conducted to determine whether posttraumatic moderate hypotension (50 to 60 mm Hg) alone would alter the histopathologic consequences of TBI. Although posttraumatic hypotension has been reported to alter the hemodynamic consequences of TBI, few studies have determined whether regional hemodynamic perturbations influence regional histopathologic outcome.

MATERIALS AND METHODS

Animal groups

Forty-five male Sprague-Dawley rats (254 to 342 g) were housed individually under controlled environmental conditions (12 hours light/12 hours dark cycle) with food and water available ad libitum before being used in the experiments. All animal procedures followed the National Institutes of Health Guide for the Care and Use of Laboratory Animals (National Institutes of Health Publication 86–23, revised 1985).

Fluid–percussion traumatic brain injury procedure

Animals were anesthetized with equithesin (a mixture of pentobarbital, propylene glycol, ethanol, MgSO₄, and chloral hydrate) 24 hours before injury and were surgically prepared for FP injury as described previously (Bramlett et al., 1999). Briefly, a craniectomy (4.8 mm in diameter) was performed over the right parietal cortex (3.8 mm posterior to the bregma and 2.5 mm lateral to the midline), leaving the dura intact. The plastic injury tube (2.6-mm inner diameter, approximately 4.8-mm outer diameter) was positioned over the exposed dura and bonded by adhesive. The tube then was cemented to the skull with dental acrylic. The scalp was sutured closed and the animals were returned to their home cages and allowed to recover.

After fasting overnight, a FP device was used to produce experimental TBI through the injury tube (Dixon et al., 1987). Intubated anesthetized rats (0.5% halothane, 30% oxygen, and 70% nitrous oxide) were subjected to a pressure pulse of moderate (1.94 to 2.18 atm) intensity. Before FP injury, catheters (PE50) were inserted into both femoral arteries for blood sampling, measurement of blood pressure, and withdrawal of reference blood samples. A silastic catheter was inserted into the femoral vein and was used to induce hypotension by exsanguination into a syringe. The right axillary artery was cannulated and the catheter (PE10) was advanced into the left ventricle for injection of microspheres. Brain temperature was monitored indirectly through a thermocouple probe inserted into the temporal muscle and maintained at 37.0°C ± 0.5°C. Rectal temperature also was monitored and maintained at 37.0°C ± 0.5°C.

Secondary hypotension

After moderate FP injury, animals were maintained for 30 minutes at either a normotensive (group 1: TBI only, n = 6) or hypotensive (group 2: TBI + hypotension, n = 6) level to simulate secondary hypotension. Hypotension (mean arterial blood pressure = 60 mm Hg) was induced by the gradual withdrawal of blood into a heparinized syringe 5 minutes after TBI. After 30 minutes of hypotension, shed blood was reinfused to restore arterial blood pressure. Blood gas measurements were taken before TBI and 10, 25, and 55 minutes after injury. Sham animals underwent all surgical procedures, including a similar period of hypotension (group 3: sham + hypotension, n = 7), but were not subjected to the FP pulse. For hemodynamic studies, an additional normotensive control group (group 4: n = 4) was produced. These animals did not undergo sham procedures.

Regional cerebral blood flow measurement

Radiolabeled microspheres (15 ± 0.5 μm in diameter; New England Nuclear, Wilmington, DE, U.S.A.) were injected into the left ventricle for measurement of rCBF. Three of 5 possible isotopes—¹⁴¹Ce, ¹¹⁴In, ¹⁰³Ru, ⁹⁵Nb, and ⁴⁶Sc—were randomly selected for injection into each animal at baseline, 20 minutes after TBI during hypotension, and 60 minutes after TBI after reinfusion of shed blood. The method for measurement of rCBF in rats has been previously described (Yamakami and McIntosh, 1989). Reference sample method for cardiac output and regional organ blood flow determinations in the rat also has been reported (Malik et al., 1976).

Approximately 125,000 microspheres were injected at each time point. Injecting this number of microspheres ensured that each brain tissue sample contained more than 400 microspheres. Beginning 10 seconds before microsphere injections, reference blood was withdrawn from the femoral artery at a rate of 0.5 mL/minute during normotension and 0.3 mL/minute during hypotension. Withdrawal continued for 45 seconds after each injection. Reference blood was replaced with 0.9% saline (1:3) after each injection. A postmortem examination was performed after each experiment to confirm position of the left ventricular catheter.

The brain was removed and fixed in 10% buffered formalin for 24 hours and then dissected into 100- to 200-g sections. Briefly, a 3-mm-thick coronal brain slice was first made at the bregma levels 1.2 mm and −1.8 mm and included cortical and subcortical areas anterior to the histopathologically vulnerable posterior cortical and subcortical regions (that is, −1.8 mm and −4.8 mm) using rodent brain matrices (Harvard Apparatus, MA, U.S.A.). The ipsilateral anterior cortical and subcortical regions adjacent to the injury were labeled as A1 and A2, respectively. The ipsilateral posterior cortical and subcortical sites that included histopathologically vulnerable regions (that is, posterior cortical and subcortical segments) were dissected and labeled as P1 and P2, respectively. Homotopic cortical and subcortical regions of the contralateral hemisphere also were dissected and labeled (that is, A3, A4, P3, P4). Radioactivity of each sample of blood and tissue was counted in a gamma counter (Auto-Gamma 5000; Packard Instruments, Meriden, CT, U.S.A.). Tissue blood flow was calculated by the standard simultaneous equation technique, and rCBF was converted to mL min⁻¹ 100 g⁻¹. Blood flow data are expressed as percent of baseline (group 4, mean ± SD).

Histopathology

Animals were killed at 72 hours after TBI (group 5: TBI only, n = 8; group 6: TBI + hypotension, n = 7) or sham surgery (group 7: sham + hypotension, n = 7). Semiserial sections were processed and stained for hematoxylin and eosin. Animals were anesthetized and perfused transcardially with isotonic saline at a pressure of 100 to 120 mm Hg for 60 seconds. This was followed by fixative for 20 minutes (FAM, a mixture of 40% formaldehyde, glacial acetic acid, and methanol; 1:1:8 by volume). After the perfusions, the heads were immersed in FAM for 24 hours. Brains then were blocked and embedded in paraffin for tissue sectioning. Coronal sections at various levels (0.8, 1.8, 3.3, 4.3, 5.8, 6.8, 7.3 mm posterior to bregma) were used for contusion area measurements. Contusion boundaries were demarcated by swollen axons, necrotic neurons, reactive astrocytes, and inflammatory cells. The cortical contusion was first traced at a power of 100× using a camera lucida microscope attachment. Contusion areas were calculated by retracing these drawings onto a digitizing table that was interfaced with a computer. Contusion volumes then were computed by using numeric integration of successive areas.

Statistical analysis

Physiologic variables and histopathologic data are expressed as mean ± SD. Group differences in contusion volume measurements and rCBF values were assessed using one-way analysis of variance.

RESULTS

Physiologic changes

Physiologic changes are summarized in Tables 1 and 2. In groups 1 and 2, mean arterial blood pressure (MABP) transiently increased at the moment of the impact, but returned to baseline values within 1 minute. Values for Paco₂, Pao₂, and pH did not differ among the experimental groups. In groups 2 and 3, MABP was significantly decreased compared with group 1 and was maintained at 59.7 mm Hg (59.7 ± 2.9 and 60.0 ± 1.3 mm Hg, respectively) during the 30-minute hypotensive period.

TABLE 1.

Physiologic parameters for CBF studies

		Postinjury
Variables	Preinjury	20 minutes	1 hour
TBI only (n = 6)
MABP	119.5 ± 5.0	114.5 ± 6.2	115.3 ± 9.5
Rectal temperature	37.0 ± 0.4	37.2 ± 0.3	37.0 ± 0.2
Brain temperature	36.7 ± 0.1	36.7 ± 0.2	36.8 ± 0.2
pH	7.449 ± 0.035	7.427 ± 0.014	7.409 ± 0.027
PaCO₂	38.0 ± 1.2	37.5 ± 1.0	37.8 ± 1.2
PaO₂	139.8 ± 15.5	140.8 ± 13.3	148.7 ± 13.6
Hct	41.7 ± 2.1		40.5 ± 3.5
TBI + hypotension (n = 6)
MABP	118.3 ± 6.9	59.7 ± 2.9*	118.5 ± 10.7
Rectal temperature	37.1 ± 0.3	36.8 ± 0.4	37.2 ± 0.2
Brain temperature	36.7 ± 0.1	36.7 ± 0.1	36.7 ± 0.2
pH	7.429 ± 0.023	7.381 ± 0.039	7.378 ± 0.077
PaCO₂	38.2 ± 1.7	39.4 ± 1.6	36.5 ± 1.3
PaO₂	136.5 ± 6.3	146.1 ± 13.4	137.7 ± 20.7
Hct	42.5 ± 2.1		40.7 ± 2.9
Sham + hypotension (n=7)
MABP	121.7 ± 6.8	60.0 ± 1.3*	123.0 ± 8.3
Rectal temperature	36.7 ± 0.1	36.9 ± 0.2	37.2 ± 0.1
Brain temperature	36.8 ± 0.1	36.8 ± 0.2	36.9 ± 0.2
pH	7.467 ± 0.017	7.391 ± 0.036	7.380 ± 0.052
PaCO₂	36.8 ± 1.1	37.9 ± 2.1	37.7 ± 1.3
PaO₂	130.8 ± 14.0	138.8 ± 16.1	137.7 ± 11.2
Hct	42.7 ± 4.6		39.8 ± 4.6
Sham (n = 4)
MABP	123.8 ± 10.2	115.5 ± 7.5	122.5 ± 8.2
Rectal temperature	36.9 ± 0.4	36.9 ± 0.1	36.9 ± 0.1
Brain temperature	36.8 ± 0.2	37.0 ± 0.2	36.9 ± 0.2
pH	7.499 ± 0.022	7.422 ± 0.033	7.386 ± 0.031
PaCO₂	38.4 ± 0.9	37.9 ± 1.8	38.7 ± 1.1
PaO₂	141.5 ± 25.3	130.8 ± 18.3	132.8 ± 11.1
Hct	43.3 ± 4.4		43.0 ± 2.9

CBF, cerebral blood flow; TBI, traumatic brain injury; MABP, mean arterial blood pressure.

P< 0.05 postinjury versus preinjury.

TABLE 2.

Physiologic parameters for histopathologic studies

		Postinjury
Variables	Preinjury	20 minutes	1 hour
TBI only (n =8)
MABP	121.9 ± 10.3	120.6 ± 10.8	123.1 ± 11.6
Rectal temperature	37.0 ± 0.4	37.2 ± 0.2	37.0 ± 0.3
Brain temperature	36.9 ± 0.2	36.8 ± 0.2	36.8 ± 0.2
pH	7.463 ± 0.039	7.449 ± 0.035	7.430 ± 0.034
PaCO₂	37.3 ± 1.4	37.8 ± 1.8	37.9 ± 1.4
PaO₂	150.0 ± 16.7	136.7 ± 11.7	133.8 ± 18.0
TBI + hypotension (n =7)
MABP	114.3 ± 6.7	59.3 ± 4.5*	112.9 ± 9.1
Rectal temperature	36.9 ± 0.2	37.3 ± 0.3	37.2 ± 0.2
Brain temperature	36.9 ± 0.2	36.8 ± 0.2	36.7 ± 0.2
pH	7.464 ± 0.044	7.424 ± 0.046	7.422 ± 0.050
paCO₂	37.8 ± 1.9	35.5 ± 2.4	38.0 ± 1.9
PaO₂	144.7 ± 12.6	131.1 ± 21.4	133.5 ± 12.7
Sham + hypotension (n =7)
MABP	114.3 ± 7.9	60.7 ± 3.5*	110.7 ± 9.3
Rectal temperature	37.1 ± 0.5	37.2 ± 0.6	37.3 ± 0.2
Brain temperature	36.9 ± 0.2	36.8 ± 0.2	36.7 ± 0.2
pH	7.436 ± 0.013	7.408 ± 0.024	7.404 ± 0.025
PaCO₂	38.4 ± 1.4	36.9 ± 1.9	38.7 ± 2.1
PaO₂	141.5 ± 20.6	132.6 ± 18.0	127.0 ± 14.8

TBI, traumatic brain injury; MABP, mean arterial blood pressure.

P< 0.05 postinjury versus preinjury.

Regional cerebral blood flow

There were no significant differences in rCBF among groups at baseline and no significant changes over time in normotensive controls. Twenty minutes after normotensive TBI, blood flow was significantly reduced in total forebrain (58% ± 12% of baseline, P < 0.01) and ipsilateral (50% ± 15%, P < 0.01) and contralateral cerebral hemispheres (65% ± 17%, P < 0.05; Fig. 1A to 1C). Hypotension further reduced blood flow 20 minutes after TBI in total forebrain (36% ± 17%, P < 0.03) and contralateral (39% ± 22%, P < 0.05), but not ipsilateral, cerebral hemispheres (33% ± 15%, P < 0.07), although there was a trend for the latter also to decrease with hypotension. Sixty minutes after TBI, blood flow was not significantly different from control in total forebrain and the contralateral cerebral hemisphere; however, in the ipsilateral hemisphere, blood flow remained less than control in both TBI groups (Fig. 1).

FIG. 1.

Values for cerebral blood flow (CBF) of total brain (A), right brain (B), left brain (C), and posterior cortex (D) expressed as percent of baseline (mean ± SD) in time controls (solid bars), hypotensive sham controls (diagonal bars), normotensive traumatic brain injury (TBI) (shaded bars), and hypotensive TBI (horizontal bars). * P < 0.05, †P < 0.01 compared with time controls or between TBI groups 20 minutes after TBI (connecting lines).

Within the histopathologically vulnerable parietal cortex (P1), induced hypotension significantly reduced rCBF 20 minutes after TBI compared with normotensive TBI values (21% ± 10% vs. 43% ± 15% of control, P < 0.01; Fig. 1D). Sixty minutes after TBI, rCBF remained less than control in normotensive TBI rats (46 ± 23, P < 0.05) but only tended to be less in hypotensive TBI rats (51% ± 34%, P = 0.09). Analysis of rCBF in other regions revealed that 20 minutes after TBI, the hypotensive group had 7 of 8 regions with rCBF less than control, whereas in the normotensive group, only 5 of 8 regions were significantly reduced (Table 3). Sixty minutes after normotensive TBI, rCBF remained less than control primarily in ipsilateral regions, whereas after reinfusion of shed blood after hypotensive TBI, rCBF was less than control in only the parietal cortex anterior to the impact (Table 3).

TABLE 3.

Regional CBF values as % of baseline (mean ± SD) 20 and 60 minutes after TBI

Variables	Hemisphere	Anterior cortex	Anterior subcortex	Posterior cortex	Posterior subcortex
20 minutes after TBI
Normotensive	R	46 ± 18†	64 ± 25	43 ± 14†	52 ± 25*
	L	57 ± 17*	72 ± 30	59 ± 22	70 ± 29*
Hypotensive	R	29 ± 13†	43 ± 20†	21 ± 10†	45 ± 18†
	L	38 ± 20†	43 ± 22†	36 ± 27	48 ± 22†
60 minutes after TBI
Normotensive	R	48 ± 17†	71 ± 25*	46 ± 22*	64 ± 27*
	L	71 ± 20	82 ± 24	66 ± 24	74 ± 22*
Hypotensive	R	47 ± 22†	71 ± 26	51 ± 34	69 ± 41
	L	59 ± 38	70 ± 37	60 ± 49	103 ± 80

CBF, cerebral blood flow; TBI, traumatic brain injury.

P < 0.05

†

P< 0.01, compared with time controls.

Histopathologic outcome

Group 7 (sham + hypotension) did not exhibit any type of cortical or subcortical damage. As seen in Fig. 2, both group 5 and 6 animals contained a well-demarcated intracerebral contusion overlying the right external capsule (Fig. 2A and 2B). However, group 6 animals showed more widespread cortical damage and a larger contusion compared with group 5. Overlying the contusion, a high frequency of necrotic cortical neurons were seen in the lateral cerebral cortex (Fig. 2C to 2F). Also, FP hypotensive rats showed more extensive cortical neuronal damage compared with normotensive rats. With hypotension, TBI rats demonstrated large numbers of shrunken eosinophilic cortical neurons in an edematous-appearing neuropil (Fig. 2F).

FIG. 2.

Photomicrographs displaying paraffin-embedded sections of rat brains three days after fluid-percussion (FP) injury. Hematoxylin and eosin staining. (A) Small contusion located at the gray-white interface in a FP normotensive rat. (B) Large contusion in a FP hypotensive rat. (C) Higher magnification of overlying cerebral cortex of FP normotensive rat showing intact neurophil with scattered damaged neurons. (D) Extensive cortical neuronal damage in FP hypotensive rat. (E) Shrunken and eosinophilic cortical neurons with scattered normal appearing neurons (arrowheads) in FP normotensive rat. (F) Few normal appearing neurons are present in edematous appearing cortical region of FP hypotensive rat. Magnification ×120 (A, B), ×300 (C, D), ×1200 (E, F).

Figure 3A shows the contusion areas at the different bregma levels in normotensive and hypotensive rats. Significant differences in contusion areas were seen at several bregma levels. Figure 3B shows overall contusion volumes. There was a significant difference in contusion volume between group 5 and 6 (0.96 ± 0.46 mm³ vs. 2.02 ± 1.25 mm³, P < 0.05).

FIG. 3.

(A) Bar graph demonstrating areas of contusion (mean ± SD) at six bregma levels in normotensive and hypotensive rats. Posttraumatic hypotension significantly increased contusion area (*) at three bregma levels (P < 0.05). (B) Contusion volume significantly increased (*) with posttraumatic hypotension (P < 0.05). TBI, traumatic brain injury.

DISCUSSION

The current study demonstrated that a mild hypotensive event after moderate FP injury significantly affects the hemodynamic and histopathologic consequences of this traumatic insult. Although previous studies have reported the effects of mild hypotension on rCBF after TBI (DeWitt et al., 1992a, 1992b; Armstead, 1999; Kroppeenstedt et al., 1999; Lewelt et al., 1980; Schmoker et al., 1992), limited data exists regarding histopathologic correlates of posttraumatic hypotension. In the current study, contusion areas and volumes were significantly increased in hypotensive FP rats compared with normothermic animals. Increased structural damage was associated with more severe hypotension-induced rCBF changes that reached ischemic levels. The current data support previous findings and extend this investigative field by showing aggravated histopathologic outcome with induced hypotension. Posttraumatic blood pressure remains an important physiologic variable in experimental and clinical TBI studies.

Measurement of rCBF revealed that normotensive TBI caused widespread reductions in rCBF in the cerebral cortex and subcortex bilaterally 20 minutes after injury. During induced hypotension, the number of regions with significant reductions in rCBF increased, and in the cerebral cortex, a further decrease in rCBF occurred within the histopathologically vulnerable PI region. These findings demonstrate the well-known loss of autoregulation after TBI and suggest that secondary ischemic injury may have caused the enhancement of the histopathologic injury seen in the hypotensive TBI group (Armstead, 1999; DeWitt et al., 1992b; Golding et al., 1999). The return of rCBF to values not different from control after reinfusion of shed blood and restoration of normal blood pressure proves that with greater perfusion pressure rCBF can be increased in the traumatized brain, and that persistent depression of rCBF did not occur after a brief period of hypotension.

The authors have previously reported that a 30-minute period of posttraumatic hypoxia exacerbates cortical and hippocampal damage after moderate FP brain injury (Bramlett et al., 1999). However, when animals were made hypoxic after TBI, mild hypotension occurred during the hypoxic period. Although hypotension is often associated with an hypoxic insult (Chesnut et al., 1993), the question of whether decreased O₂ or moderate hypotension was primarily responsible for the exacerbation of brain damage in that study remains unclear. The current results indicate that the hypotensive component of the hypoxic insult most likely had a significant impact on tissue injury. However, because the histopathologic consequences were more severe with secondary hypoxia compared with hypotension alone (Bramlett et al., 1999), the authors conclude that a transient period of reduced Po₂ alone has a detrimental effect on traumatic outcome.

Kroppenstedt and colleagues (1999) reported that a reduction of MABP to 70 mm Hg significantly increased contusion volumes after cortical impact injury. Interestingly, the range of mean arterial pressure with no enlargement of contusion volume was between 70 and 105 mm Hg. Furthermore, a brief period of 30-minute hypotension was enough to cause a significant increase in contusion volume. Other studies have reported the effects of hypotension after TBI in a weight drop model, cryogenic model, and FP model (DeWitt et al., 1992a, 1992b; Giri et al., 2000; Golding et al., 1999; Lewelt et al., 1980). In each case, hypotension appeared to adversely affect the hemodynamic, electrophysiologic, or metabolic state of the traumatized brain. Because MABP of 60 mm Hg is apparently an important threshold for secondary injuries in rats, it may be important in future studies directed at mechanistic issues to also investigate more severe degrees of hypotension.

As previously reported, moderate parasagittal FP injury produces a well-demarcated contusion, primarily overlying the lateral external capsule, with damaged cortical neurons within the overlying parietal cortex (Bramlett et al., 1999; Dietrich et al, 1994, 1996b). This relatively focal lesion is a consequence of mechanically disrupted blood vessels because of shearing stress caused by FP injury (Cortez et al., 1989; Dietrich et al., 1994). Blood–brain barrier breakdown and subsequent reductions in local CBF (lCBF) occur in the damaged region (Cortez et al., 1989; Dietrich et al., 1994, 1996a; Yamakami and McIntosh, 1991). In the current study, induced hypotension increased overall contusion volume in the lateral parietal cortex. Because posttraumatic hypotension also aggravated the hemodynamic consequences of this specific brain region, vascular mechanisms most likely participated in the observed outcome.

In this regard, Barzo et al. (1996) reported that posttraumatic secondary insults after closed head injury prolonged the time of blood–brain barrier breakdown. Perturbations in blood–brain barrier function possibly leading to increased brain edema (Baldwin et al., 1996), coupled with induced hypotension, might lead to more severe hemodynamic disturbances. Such a consequence would be expected to cause additional brain damage, possibly by ischemic mechanisms. In this study, induced hypotension led to rCBF values less than 18 mL 100 g⁻¹ min⁻¹, which are considered ischemic in rodents (Bolander et al., 1989; Tyson et al., 1984). However, because of the complex metabolic consequences of posttraumatic brain, ischemic thresholds in normal brain may not directly apply to the posttraumatic brain (Giri et al., 2000). The regional measurement of adenosine triphosphate levels and other energy metabolites would also provide important information regarding the metabolic status of the posttraumatic brain after secondary hypotensive insult (Ishige et al., 1988).

Previous studies have demonstrated that metabolic rates are initially increased after TBI, with decreased rates observed at later study periods (Yoshino et al., 1991). Clinical and experimental data also have indicated that flow–metabolic uncoupling can occur in vulnerable brain regions after TBI and may complicate the interpretation of the significance of posttraumatic hemodynamic alterations (Ginsberg et al., 1997; Jaggi et al., 1990). In this regard, studies in which measurements of cerebral oxygenation have been undertaken have provided important data about the relative adequacy of CBF after brain injury. Cryogenic injury combined with reduction of mean arterial blood pressure of 50 mm Hg/45 minutes was reported to significantly decrease cerebral oxygen delivery and cerebral metabolic rate of oxygen (Lewelt et al., 1980). In a recent study, Giri and colleagues (2000) reported, using a cortical impact injury model, that brain tissue Po₂ and rCBF decreased to low levels in the posttraumatic brain during a secondary carotid occlusion insult; thus, reduced CBF in that study was inadequate to prevent oxygen depletion. Similar studies are required in the current FP model to determine whether metabolic requirements are significantly influenced under induced hypotensive periods.

Mechanisms underlying the autoregulatory dysfunction after TBI require clarification (Golding et al., 1999). Traumatic brain injury is known to produce a marked increase in vasoactive substances, including prostaglandins (Armstead, 1998; DeWitt et al., 1997; Ellis et al., 1981) and oxygen free radicals (Kontos and Wei, 1986), that may affect rCBF and vascular reactivity. Several studies have demonstrated that pial arteries and arterioles do not exhibit normal vasodilatory responses to acetylcholine and serotonin after TBI (Kontos and Wei, 1992). Armstead (1999) measured marked elevations in cerebrospinal fluid concentrations of endothelin-1 after TBI in piglets and reported that pretreatment with an endothelin-1 antagonist partially restored pial artery dilation during hypotension. DeWitt and colleagues (1997) reported that treatment with superoxide dismutase and L-arginine prevented or reversed posttraumatic hypoperfusion after midline FP in rats. Thus, early treatment with agents that target free radical generation, nitric oxide production, or vasoactive substances may promote protection from secondary insults.

In summary, posttraumatic hypotension after FP injury significantly worsens histopathologic outcome in an established FP model of TBI. It is concluded that hypoxia-induced hypotension may have contributed to previous results regarding the detrimental consequences of posttraumatic hypoxia after TBI. However, because hypotension appears to have a milder histopathologic effect on outcome compared with hypoxia, critical levels of O₂ delivery after TBI also appear to be a critical factor in secondary injury mechanisms. Strategies to prevent or reverse autoregulatory abnormalities and enhance normal O₂ delivery to posttraumatic tissues offers a reasonable approach to preventing acute as well as progressive damage after TBI.

Footnotes

Acknowledgments:

The authors gratefully acknowledge the superb technical assistance of Susan Kraydieh and Charlaine Rowlette.

References

Adams

Gennarelli

Graham

(1982) Brain damage in non-missile head injury: Observations in man and subhuman primates. In: Recent advances in neuropathology ( Smith

Cavanagh

, eds), Edinburgh: Churchill Livingstone, pp 165–190

Armstead

Kurth

(1994) Different cerebral hemodynamic responses following fluid percussion brain injury in the newborn and juvenile pig. J Neurotrauma 111:487–497

Armstead

(1998) Brain injury impairs prostaglandin cerebrovasodilation. J Neurotrauma 15:721–729

Armstead

(1999) Role of endothelin-1 in age-dependent cerebrovascular hypotensive responses after brain injury. Am J Physiol H1884–H1894

Baldwin

Fugaccia

Brown

Scheff

(1996) Blood-brain barrier breach following cortical contusion in the rat. J Neurosurg 85:476–481

Barzo

Marmarou

Fatouros

Corwin

Dunbar

(1996) Magnetic resonance imaging-monitored acute blood-brain barrier changes in experimental traumatic brain injury. J Neurosurg 85:1113–1121

Bolander

Persson

Hillered

D'Argy

Ponten

Olsson

(1989) Regional cerebral blood flow and histopathologic changes after middle cerebral artery occlusion in rats. Stroke 20:930–937

Bouma

Muizelaar

(1992) Cerebral blood flow, cerebral blood volume, and cerebrovascular reactivity after severe head injury. J Neurotrauma 9(suppl 1):S333–S348

Bramlett

Green

Dietrich

(1999) Exacerbation of cortical and hippocampal CA1 damage due to posttraumatic hypoxia following moderate fluid-percussion brain injury in rats. J Neurosurg 91:653–659

10.

Bryan

Cherian

Robertson

(1995) Regional cerebral blood flow after controlled cortical impact injury in rats. Anesth Analg 80:687–695

11.

Cherian

Robertson

Goodman

(1996) Secondary insults increase injury after controlled cortical impact in rats. J Neurotrauma 13:371–383

12.

Chesnut

Marshall

Klauber

Blunt

Baldwin

Eisenberg

Jane

Marmarou

Foulkes

(1993) The role of secondary brain injury in determining outcome from severe head injury. J Trauma 34:216–222

13.

Clark

RSB

Kochanek

Dixon

(1997) Early neuropathologic effects of mild or moderate hypoxemia after controlled cortical impact injury in rats. J Neurotrauma 14:179–189

14.

Cortez

McIntosh

Oble

(1989) Experimental fluid percussion brain injury: Vascular disruption and neuronal and glial alterations. Brain Res 482:271–282

15.

Davis

Jenkins

DeWitt

Prough

(1998) Mild traumatic brain injury does not modify the cerebral blood flow profile of secondary forebrain ischemia in Wistar rats. J Neurotrauma 15:615–625

16.

DeWitt

Jenkins

Wei

(1986) Effects of fluid-percussion brain injury on regional cerebral blood flow and pial arteriolar diameter. J Neurosurg 64:787–794

17.

DeWitt

Prough

Taylor

Whitley

(1992a) Reduced cerebral blood flow, oxygen delivery, and electroencephalographic activity after traumatic brain injury and mild hemorrhage in cats. J Neurosurg 76:812–821

18.

DeWitt

Prough

Taylor

Whitley

Deal

Vines

(1992b) Regional cerebrovascular responses to progressive hypotension after traumatic brain injury in cats. Am J Physiol 263:H1276–H1284

19.

DeWitt

Jenkins

Prough

(1995) Enhanced vulnerability to secondary ischemic insults after experimental traumatic brain injury. New Horizons 3376–383

20.

DeWitt

Smith

Deyo

Miller

Uchida

Prough

(1997) L-arginine and superoxide dismutase prevent or reverse cerebral hypoperfusion after fluid-percussion traumatic brain injury. J Neurotrauma 14:223–233

21.

Dietrich

Alonso

Halley

(1994) Early microvascular and neuronal consequences of traumatic brain injury. A light and electron microscopic study in rats. J Neurotrauma 11289–11301

22.

Dietrich

Alonso

Busto

Prado

Dewanjee

Ginsberg

(1996a) Widespread hemodynamic depression and focal platelet accumulation after fluid percussion brain injury: A double-label autoradiographic study in rats. J Cereb Blood Flow Metab 16:481–489

23.

Dietrich

Alonso

Halley

Busto

(1996b) Delayed posttraumatic brain hyperthermia worsens outcome after fluid percussion brain injury: A light and electron microscopic study in rats. Neurosurgery 38:533–541

24.

Dietrich

Alonso

Busto

Prado

Zhao

Dewanjee

Ginsberg

(1998) Posttraumatic cerebral ischemia after fluid percussion brain injury: An autoradiographic and histopathological study in rats. Neurosurgery 43:585–593

25.

Dixon

Lyeth

Povlishock

Findling

Hamm

Marmarou

Young

Hayes

(1987) A fluid percussion model of experimental brain injury in the rat. J Neurosurg 67:110–119

26.

Ellis

Wright

Wei

Kontos

(1981) Cyclo-oxygenase products of arachidonic acid metabolism in cat cerebral cortex after experimental concussive brain injury. J Neurochem 37:892–896

27.

Engleborghs

Haseldonckx

Van Reempts

Van Rossem

Wouthers

Borgers

Verlooy

(2000) Impaired autoregulation of cerebral blood flow in an experimental model of traumatic brain injury. J Neurotrauma 17:667–677

28.

Ginsberg

Zhao

Alonso

Loor-Estades

Dietrich

Busto

(1997) Uncoupling of local cerebral glucose metabolism and blood flow after acute fluid-percussion injury in rats. Am J Physiol 272:H2859–H2868

29.

Giri

Krishnappa

Bryan

Jr Robertson

(2000) Regional cerebral blood flow after cortical impact injury complicated by a secondary insult in rats. Stroke 31:961–967

30.

Golding

Contant

Jr Robertson

Bryan

Jr (1998) Temporal effect of severe controlled cortical impact injury in the rat on the myogenic response of the middle cerebral artery. J Neurotrauma 15:973–984

31.

Golding

Robertson

Bryan

Jr (1999) The consequences of traumatic brain injury on cerebral blood flow and autoregulation: A review. Clin Exp Hypertens 21:299–332

32.

Graham

Adams

Doyle

(1978) Ischemic brain damage in fatal non-missile head injury. J Neurol Sci 39:213–234

33.

Ishige

Pitts

Berry

(1987) The effect of hypoxia on traumatic head injury in rats: Alterations in neurologic function, brain edema, and cerebral blood flow. J Cereb Blood Flow Metab 7:759–767

34.

Ishige

Pitts

Berry

Nishimura

James

(1988) The effects of hypovolemic hypotension on high-energy phosphate metabolism of traumatized brain in rats. J Neurosurg 68:129–136

35.

Jaggi

Obrist

Gennarelli

(1990) Relationship of early cerebral blood flow and metabolism to outcome in acute head injury. J Neurosurg 72:176–182

36.

Jenkins

Moszynski

Lyeth

Lewelt

DeWitt

Allen

Dixon

Povlishock

Majewski

Clifton

(1989) Increased vulnerability of the mildly traumatized rat brain to cerebral ischemia: The use of controlled secondary ischemia as a research tool to identify common or different mechanisms contributing to mechanical and ischemic brain injury. Brain Res 477:211–224

37.

Kelly

Martin

Kordestani

(1997) Cerebral blood flow as a predictor of outcome following traumatic brain injury. J Neurosurg 86:633–641

38.

Kochanek

Marion

Zhang

Shiding

White

Palmer

Clark

RSB

O'Malley

Styren

DeKosky

(1995) Severe controlled cortical impact in rats. Assessment of cerebral edema, blood flow, and contusion volume. J Neurotrauma 12:1015–1025

39.

Kontos

Wei

(1986) Superoxide production in experimental brain injury. J Neurosurg 64:803–807

40.

Kontos

Wei

(1992) Endothelium-dependent responses after experimental brain injury. J Neurotrauma 9:349–354

41.

Kroppenstedt

Kern

Thomale

Schneider

Lanksch

Unterberg

(1999) Effect of cerebral perfusion pressure on contusion volume following impact injury. J Neurosurg 90:520–526

42.

Law

Hovda

Cryer

(1996) Fluid percussion brain injury adversely affects control of vascular tone during hemorrhagic shock. Shock 6:213–217

43.

Lewelt

Jenkins

Miller

(1980) Autoregulation of cerebral blood flow after experimental fluid percussion injury of the brain. J Neurosurg 53:500–511

44.

Malik

Kaplan

Saba

(1976) Reference sample method for cardiac output and regional blood flow determinations in the rat. J Appl Physiol 40:472–475

45.

Marion

Darby

Yonas

(1991) Acute regional cerebral blood flow changes caused by severe head injuries. J Neurosurg 74:407–414

46.

Martin

Patwardhan

Alexander

(1997) Characterization of cerebral hemodynamic phases following severe head trauma: Hypoperfusion, hyperemia, and vasospasm. J Neurosurg 87:9–19

47.

Mathew

DeWitt

Bryan

Jr Bukoski

Prough

(1999) Traumatic brain injury reduces myogenic responses in pressurized rodent middle cerebral arteries. J Neurotrauma 16:1177–1186

48.

Muizelaar

Lutz

Becker

(1984) Effect of mannitol on ICP and CBF and correlation with pressure autoregulation in severely head injured patients. J Neurosurg 9:700–706

49.

Nawashiro

Shima

Chigasaki

(1995) Selective vulnerability of hippocampal CA3 neurons to hypoxia after mild concussion in the rat. Neurol Res 17:455–460

50.

Newell

Weber

Watson

Aaslid

Winn

(1996) Effect of transient moderate hyperventilation on dynamic cerebral autoregulation after severe head injury. Neurosurgery 39:35–43

51.

Nilsson

Gazelius

Carlson

Hillered

(1996) Continuous measurement of changes in regional cerebral blood flow following cortical compression contusion trauma in the rat. J Neurotrauma 13:201–207

52.

Obrist

Langfitt

Jaggi

(1984) Cerebral blood flow and metabolism in comatose patients with acute head injury. Relationship to intracranial hypertension. J Neurosurg 61:241–253

53.

Povlishock

Becker

Sullivan

Miller

(1978) Vascular permeability alterations to horseradish peroxidase in experimental brain injury. Brain Res 153:223–239

54.

Schmoker

Zhuang

Shackford

(1992) Hemorrhagic hypotension after brain injury causes an early and sustained reduction in cerebral oxygen delivery despite normalization of systemic oxygen delivery. J Trauma 32:714–720

55.

Tyson

Teasdale

Graham

McCulloch

(1984) Focal cerebral ischemia in the rat: Topography of hemodynamic and histopathological changes. Ann Neurol 15:559–567

56.

Wei

Dietrich

Povlishock

Navari

Kontos

(1980) Functional, morphological, and metabolic abnormalities of the cerebral microcirculation after concussive brain injury in cats. Circ Res 46:37–47

57.

Yamakami

McIntosh

(1991) Alterations in regional cerebral blood flow following brain injury in the rat. J Cereb Blood Flow Metab 11:655–660

58.

Yoshino

Hovda

Kawamata

(1991) Dynamic changes in local cerebral glucose utilization following cerebral concussion in rats: Evidence of a hyper- and subsequent hypometabolic state. Brain Res 561:106–119

Delayed Hemorrhagic Hypotension Exacerbates the Hemodynamic and Histopathologic Consequences of Traumatic Brain Injury in Rats

Abstract

Keywords

MATERIALS AND METHODS

Animal groups

Fluid–percussion traumatic brain injury procedure

Secondary hypotension

Regional cerebral blood flow measurement

Histopathology

Statistical analysis

RESULTS

Physiologic changes

Regional cerebral blood flow

Histopathologic outcome

DISCUSSION

Footnotes

Acknowledgments:

References