INTRODUCTION

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THE "PUSH-PULL" EFFECT (3) denotes the reduced tolerance to +G_z (hypergravity) when +G_z stress is preceded by exposure to hypogravity, i.e., fractional, zero, or negative G_z. The reduction in G_z tolerance likely stems from the regional changes in the hydrostatic component of blood pressure imposed by hypogravity and the sudden reversal of these changes by subsequent exposure to hypergravity. The changes in the hydrostatic pressure likely elicit the push-pull effect via their action on the mechanically sensitive, reactive characteristics of the circulation (e.g., baroreflex, myogenic, and vestibular regulatory mechanisms). The push-pull effect has been demonstrated in several studies on human subjects (2-4) and has been demonstrated to occur in male rats (18). The push-pull effect has advanced from an unappreciated flight hazard (2) to an acknowledged source of risk for G-induced loss of consciousness (15). Moreover, similar alterations in regional hydrostatic pressure occur during common everyday movements such as bending over to pick something up and thus may relate to the dizziness that can accompany "standing up too quickly."

In the present study, we sought to explore the influence on the push-pull effect of two factors that are thought to modulate responses to G_z stress. The first factor was gender. Previous studies on human subjects provide conflicting evidence as to the importance of gender on G_z tolerance. Some investigators have found that the tolerance to simulated orthostatic stress imposed by lower body negative pressure is less in female subjects (5, 17, 20), whereas others have found that gender is not a factor (7, 8). Nevertheless, gender is generally thought to exert an important influence on G_z tolerance inasmuch as the incidence of clinically relevant orthostatic hypotension is greater in female than in male subjects (14). Also, to our knowledge no studies have evaluated the influence of gender on the push-pull effect. Therefore, we sought to address the following two questions. First, does gender influence the eye-level blood pressure (ELBP) response to brief +G_z stress in rats? Second, does gender influence the push-pull effect?

The second factor we sought to evaluate was whether altering the axis of rotation about which push-pull G_z stress is imposed alters the expression of the push-pull effect. Recently Cheung et al. (4) demonstrated that arterial pressure falls further and faster during head-up tilt when a "push-pull" G_z profile is imposed by "rolling" (x-axis rotation) a subject compared with "pitching" (y-axis rotation) a subject. We sought to test whether this orientation-dependent influence seen in a relatively tall biped (humans) is also seen in a relatively short quadruped. Studies were carried out on isoflurane-anesthetized rats in which gravitational stress was imposed by tilting. Finally, because volatile anesthetics can diminish sympathetic responses (6), similar studies were repeated on five male and five female rats anesthetized with ketamine-xylazine to compare the effects of these anesthetic agents.

	METHODS

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The following procedures meet National Institutes of Health guidelines and were reviewed and approved by the Institutional Animal Care and Use Committee of the University of Iowa.

Surgical preparation. Ten male (270-300 g) and 10 female (240-300 g) Sprague-Dawley rats were anesthetized with isoflurane and restrained supine on a board. A catheter was implanted in the right carotid artery. The animal's body temperature was maintained at 37°C with a homeothermically controlled heat lamp connected to a rectal thermistor. In a separate group, five males (280-360 g) and five females (260-290 g) were anesthetized with ketamine-xylazine (91 mg/kg ketamine; 9.1 mg/kg xylazine ip) and exposed to the same tilt protocol for comparison of effects of anesthetics.

Experimental procedures. For pitch rotations, the board was oriented such that Earth's gravity vector was applied across the animal's x-axis so that G_z gravitational stress could be imposed by manually rotating the board and thus the animal ±90° about the animal's y-axis. For roll rotations, the board was oriented such that Earth's gravity vector was applied across the animal's y-axis so that G_z gravitational stress could be imposed by manually rotating the board and thus the animal ±90° about the animal's x-axis. The starting position was 0 G_z (+1 G_x or G_y). The control treatment consisted of rotating the animal 90° head up (+1 G_z) for 10 s. The push-pull treatment consisted of 10 s of head-up tilt immediately preceded by 2 s of 90° head-down tilt (1 G_z). The animal recovered for at least 2 min in the horizontal (0 G_z) position between tilts. A schematic illustration of the experimental protocol is shown in Fig. 1. Eight G profiles were imposed (2 control and 2 push-pull profiles for each of the 2 axes of rotation) in a counterbalanced design to minimized possible time effects of repeated exposure to gravitational stress. Each rat was subjected to this series of eight G profiles twice.

View larger version (10K): [in this window] [in a new window]   Fig. 1.   Schematic illustration of G profiles.

Data collection. The arterial catheter was connected to a pressure transducer (PE10 EZ, Ohmeda, Madison, WI) secured at the level of the heart along the G_z axis and just above (anterior to) the sternum when the animal was supine. Because the transducer was located above the heart with respect to gravity when the animal was in this posture (and this posture only), a correction factor of 2 mmHg was added to the measured pressure during periods when the animal was supine (+1 G_x). A length of water-filled tubing connected to a similar pressure transducer was mounted on the board to measure tilt.

Data analysis. Data analysis was performed on 1-s averages of the digitized data. Baseline ELBP was established by calculating the average pressure over the 5 s immediately preceding tilt onset. The magnitude of the ELBP response to the "push" (G_z) stress was calculated as the difference between the peak pressure during the push (taken at a time when the animal was stationary) and baseline pressure. The magnitude of the ELBP response to the "pull" (+G_z) stress was calculated as the difference between baseline ELBP and the ELBP observed at 3 s after the onset of head-up tilt.

Statistical analysis. The main effects of gender, axis of rotation, and push-pull treatment on 1) baseline ELBP, 2) the magnitude of the push, and 3) the magnitude of the pull (+G_z) stress were tested by multiple linear regression (10). Dummy variables were used as independent variables to encode for difference of gender, orientation (pitch vs. roll), and control vs. push-pull treatment. In addition, paired t-tests were used to determine whether males and females separately expressed a push-pull effect. Data are reported as means ± SE.

	RESULTS

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Isoflurane. Across the data set as a whole, there was a statistically significant push-pull effect (P < 0.01), i.e., ELBP fell further below baseline during +G_z stress when +G_z stress was immediately preceded by 2 s of G_z stress. This is apparent in Fig. 2, which shows representative data from a female rat in response to pitch rotation.

View larger version (24K): [in this window] [in a new window]   Fig. 2.   Response of eye-level blood pressure to a control +Gz gravitational stress (A) and push-pull gravitational stress (C) imposed by pitching an anesthetized female rat. B and D: degrees of the tilt during control and push-pull tilt, respectively.

Gender. The influence of gender on ELBP during control and push-pull gravitational stress is presented graphically in Fig. 3. Data from pitch and roll trials were averaged together to produce these plots. Gender exerted a statistically significant effect on baseline ELBP (P < 0.01) with pressure being ~4 mmHg higher in the females. Thereafter, the responses in the males and females essentially parallel one another (Fig. 3A). To verify this observation, the data were plotted to show the changes in ELBP relative to baseline (i.e., the baseline value of ELBP was subtracted from the value of ELBP observed at each point in time), and these curves are shown in Fig. 3B. It can be seen that the responses from males and females are essentially superimposed. The statistical analysis confirmed what is visually apparent in Fig. 3. ELBP rose to a similar extent (~8 mmHg) during push (+G_z) stress in both the males and females (Table 1). ELBP fell to a similar extent (~18 mmHg) during control +G_z stress in both males and females (Table 1). Also, ELBP fell to a similar extent (~22 mmHg) during push-pull +G_z stress in both males and females (Table 1). Finally, when analyzed separately, both males (P < 0.001) and females (P < 0.001) expressed a statistically significant push-pull effect.

View larger version (20K): [in this window] [in a new window]   Fig. 3.   Male and female rats express similar push-pull effects. A: group mean responses of eye-level blood pressure from 10 males and 10 females during control (C) and push-pull (PP) tilts. B: relative change in eye-level blood pressure from baseline. Values are means ± SE.

Pitch vs. roll. The influence of the axis of rotation on ELBP during control and push-pull gravitational stress is presented graphically in Fig. 4. Data from males and females were averaged together to produce these plots. It can be seen that little difference is observed between the pitch and the roll trials. The statistical analysis confirmed what is visually apparent in Fig. 4. The alteration in orientation of the rats with respect to Earth's gravity vector between the pitch and the roll trials did not influence baseline ELBP (Table 2). Also, ELBP rose to a similar extent (~8 mmHg) regardless of whether push (+G_z) stress was imposed by pitching or rolling the animal (Table 2). Finally, ELBP fell to a similar extent during both control and push-pull treatment regardless of the axis about which the rat was rotated.

View larger version (15K): [in this window] [in a new window]   Fig. 4.   Axis of rotation does not influence the responses of eye-level blood pressure to control and push-pull gravitational stress. Values are means ± SE; n = 20 rats.

Ketamine-xylazine. In contrast to the partial recovery of ELBP seen during the 10 s of +1 G_z tilt during isoflurane anesthesia (Figs. 3 and 4), ELBP deteriorated further over this period under ketamine-xylazine anesthesia (Fig. 5). Figure 5 also shows that ketamine-xylazine abolished the push-pull effect. Overall, responses under ketamine-xylazine were similar between males and females.

View larger version (15K): [in this window] [in a new window]   Fig. 5.   Neither males (n = 5) nor females (n = 5) exhibited a push-pull effect during ketamine-xylazine anesthesia. Values are means ± SE.

	DISCUSSION

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Isoflurane. The principal new findings of the present study are threefold. First, female rats exhibit a push-pull effect, but gender does not appear to affect the magnitude of the push-pull effect. Second, gender does not appear to influence the extent to which ELBP falls during brief +G_z stress imposed by tilting anesthetized rats. Third, pitch vs. roll rotation does not appear to influence ELBP during G_z stress imposed by tilting anesthetized rats.

+1 G_z stress. Across conditions, ELBP was partially restored toward baseline levels during 10 s of head-up tilt, thereby demonstrating that cardiovascular reflexes appear to function appropriately in this setting. Also, the magnitude of the changes in pressure we observed were as expected for an animal with a relatively short heart-to-eye distance, i.e., pressure changes would be expected to be about five times greater in humans owing to their fivefold greater heart-to-eye distance.

Push-pull. The push-pull effect, i.e., the ELBP-reducing effect of brief exposure to hypogravity immediately before +G_z stress, is observed across a wide variety of conditions and subjects. A push-pull effect is expressed in human subjects in-flight during aerial combat maneuvering (1) as well as during ground-based centrifugation studies (2, 3). It is also observed in human subjects during tilting (4) and is apparent in human subjects during parabolic flight profiles (13). A push-pull effect is also seen in conscious animals during centrifugation and in anesthetized animals during tilting (18). Thus the push-pull effect appears to constitute a stable, predictable physiological phenomenon.

Gender. Previous research on orthostatic (+G_z) tolerance in males and females provides mixed results. Some of the discrepancy may stem from the different approaches used to gauge orthostatic tolerance. For example, although a centrifuge study found G tolerances overall to be the same in men and women, there was a strong negative correlation of height and G tolerance (9). Thus, in height-matched groups of men and women, women were found to have a lower G tolerance (8). Tilt tests and stand tests constitute two additional height-dependent tests of orthostatic tolerance. A tilt-table study of human subjects revealed that young men and women have similar tilt tolerances (not height matched); gender differences in heart rate and blood pressure during tilt were attributed to differences in these variables in recumbency (19). Many investigators evaluate orthostatic tolerance by simulating G_z stress by using lower body negative pressure, a relatively height-independent measure of G tolerance. Most (5, 12, 17) but not all (7) studies report that women have a lower tolerance to lower body negative pressure.

Pitch vs. roll. A recent study by Cheung et al. (4) examined the importance of the axis of rotation on responses to G_z stress in human subjects. From a 75° head-up position, subjects were rotated to a 45° head-down (push) position and then rotated back to the 75° head-up position (pull). Tilts were carried out about the x-axis (roll) and about the y-axis (pitch). The magnitude of the fall in systolic blood pressure induced by head-up rotation in the roll position was found to be 10 mmHg greater than during the same maneuver in the pitch position (4). The lesser fall in pressure during pitch rotation was associated with a more potent tachycardia. The results of the present study indicate that the axis of rotation does not influence the push-pull effect in anesthetized rats subjected to tilting. Humans generally encounter more pitch than roll maneuvers in day-to-day activity and, on the basis of the study by Cheung et al., are better at compensating for cardiovascular consequences of this type of movement (4). Clearly there are important species and methodological differences between our study and that of Cheung et al. (e.g., in contrast to the mean pressures reported in the present study, Cheung et al. reported that systolic pressure and the response of mean pressure in their subjects could possibly differ) that could contribute to the observed differences. Another potential explanation for the discrepancies of our results and the results of Cheung and co-workers is the fact that, because +G_z stress is not nearly the threat to blood pressure regulation in rats as it is in humans, rats may be endowed with less well-organized regulatory mechanisms to combat this stress than are humans, possibly limiting their usefulness for exploring this issue.

Ketamine-xylazine. Although there are concerns about isoflurane inhibiting cardiovascular reflex responses (6) and thus potentially confounding our results, ketamine-xylazine appears to exert a more profound inhibition inasmuch as blood pressure continued to deteriorate throughout the brief period of G_z stress, whereas pressure was partially restored under isoflurane anesthesia. On the basis of these observations, isoflurane is the anesthetic of choice.