Regional distribution of brain blood flow during maximal exertion in splenectomized ponies

Respiration Physiology, 68 (1987) 77-84 Elsevier

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RSP 01261

Regional distribution of brain blood flow during maximal exertion in splenectomized ponies Murli Manohar Department of Veterinary B iosciences, Collegeof VeterinaryMedicine, Universityof Illinois at Urbana-Champaign, Urbana, IL 61801, U.S.A. (Accepted for publication 8 December 1986) Abstract. It has been reported in exercising ponies that 02 supply to all regions of the brain increased primarily due to a large increment in Cao2 and it was implied that this may reflect a generalized increase in brain metabolism during strenuous exercise. Spleneetomy ameliorates the rise in Cao2 observed with exercise in ponies. Thus, the objective of the present study was to examine changes in regional brain blood flow and 02 supplyof splenectomizedponies with sub-maximaland maximal exercise and to compare these data with previous observations in normal ponies. It was reasoned that in the absence of a marked rise in Cao2, the brain blood flow of splenectomizedponies would have to increase markedly if brain metabolism also increased with severe exercise. Regional brain blood flow was studied using 15 #m diameter radionuclide labeled microspheres injected into the left atrium during rest (control) and sub-maximal as well as maximal exertion on a treadmill. It was observed that despite marked arterial hypoeapnia and acute systemic hypertension which developed during exercise, blood flow as well as 02 supply in the cerebral cortex, caudate nuclei, cerebral white matter, cerebellar white matter, thalamus-hypothalamus, mid-brain, pons and medulla were not different from control values. In the cerebellar cortex, however, blood flow and 02 supply increased with both work intensities. Thus, it was concluded that in exercisingponies, metabolic 02 requirement increased in the cerebellar cortex but was most likely not different from control (rest) in other regions of the brain. Blood flow; Brain; Cerebral circulation; Hemodynamics; Microspheres

I n a recent study o n n o r m a l healthy ponies, it was reported that during maximal exercise blood flow in the cerebellar gray matter, pons a n d medulla increased but in other brain regions perfusion remained near baseline (rest) values despite marked arterial hypoc a p n i a a n d acute systemic hypertension that existed during exercise (Manohar, 1986). However, due to a 60~o increment in hemoglobin concentration the arterial 0 2 concentration (Cao2) increased by 58~o a n d therefore, 0 2 supply in all regions of the brain increased significantly (P < 0.01) during exercise ( M a n o h a r , 1986). Because a tight coupling of brain 0 2 d e m a n d a n d supply is k n o w n to exist (Heistad a n d K o n t o s , 1983), it was inferred that increased 0 2 supply to all areas of the p o n y brain during

Correspondence address: Dr. M. Manohar, Department of Veterinary Biosciences, 212 Large Animal Clinic, College of Veterinary Medicine, University of Illinois, Urbana, IL 61801, U.S.A. 0034-5687/87/$03.50 © 1987 Elsevier Science Publishers B.V. (Biomedical Division)

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strenuous exertion may signal an augmentation of overall brain metabolism. The objective of the present study, therefore, was to seek evidence to support or refute this hypothesis by obtaining data from splenectomized ponies during strenuous exertion because splenectomy is known to ameliorate the rise in Cao2 observed in horses during exercise (Persson and Lydin, 1973). It was reasoned that in the absence of a marked increment in Cao2 , the regional brain blood flow of splenectomized ponies would have to increase markedly if brain metabolism increased during strenuous exercise. Regional distribution of brain blood flow in the present study was examined using 15 #m diameter radionuclide labeled microspheres in nine healthy splenectomized ponies at rest and during sub-maximal and maximal exercise. Methods

Experiments were carried out on 9 healthy grade ponies of both sexes, 2-5 years old and weighing between 160 and 184 kg (170 + 4 kg; mean + SEM). The ponies were dewormed and had received tetanus toxoid several days before surgery. Splenectomy was performed on these animals 4 to 9 weeks before the hemodynamic study using general anesthesia induced with glyceryl guaiacolate + thiamyl sodium administered intravenously and maintained with 1-1.5 ~o halothane vaporized in 0 2. Our procedures for the hemodynamic studies have been described in detail previously (Parks and Manohar, 1983a,b; Manohar, 1986). All ponies were accustomed to being handled by people and were trained to run on a treadmill.

Experimental protocol It was observed that following splenectomy, the exercise capacity of the ponies was markedly diminished. Thus, about 36-48 h before the brain blood flow study, we determined the treadmill speed at which these ponies could work (at maximal cardiac frequency) for 3.75-4.00 min. Usually this varied from 14 to 15 miles per hour. Although this treadmill speed would provide maximal exercise for splenectomized ponies, it would clearly be a sub-maximal effort for normal healthy ponies. The splenectomized ponies were studied during the following conditions. Measurements were made during steady state conditions as judged by the stability of heart rate, pressures in the aorta and pulmonary artery, and arterial blood-gas tensions. (1) Rest(control/base-line). Data were obtained from splenectomized ponies standing comfortably on a treadmill as described previously (Manohar, 1986). (2) Sub-maximal exercise. The ponies were exercised on a treadmill at a speed setting equal to 75 ~ of that needed for maximal exercise in splenectomized ponies (see above). Measurements were made during the third minute when a steady state is known to exist (Bisgard et al., 1978). Exercise was discontinued at 3.75 min and a rest period of 90 rain was allowed to permit various cardio-respiratory variables to return toward pre-exercise control values.

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(3) Maximal exercise. The splenectomized ponies were exercised at a predetermined work intensity which could not be sustained for more than 4.00 min. Measurements were made during the third minute of exercise during steady state conditions (Bisgard et al., 1978). Exercise was discontinued at 3.75 min and a rest period of 90 min followed. At each step of the protocol, we determined cardiac frequency, phasic and mean aortic pressure (catheter-tip-micromanometers, Millar Instruments, Houston, TX), regional brain blood flow and 02 delivery, arterial blood-gas tensions, pHa (BMS3MK2/PHM73 Radiometer, Copenhagen, Denmark), hemoglobin concentration and hemoglobin-O 2 saturation (OSM2 Radiometer). The Cao: (ml. dl- 1; STPD) was calculated as hemoglobin concentration (g.d1-1) x 1.34 × 0 2 saturation plus 0.003 × Pao~ in mm Hg. Regional distribution of brain blood flow was examined using radionuclide labeled 15 #m diameter microspheres (Parks and Manohar, 1983a,b; Manohar, 1986). A well-agitated ultrasonicated suspension of 12-18 million microspheres, verified for lack of clumping with a microscope, was injected into the left atrium while reference arterial blood was being withdrawn from the thoracic aorta at 28.00 ml. min- 1. The sequence of various nuclides used (141Ce, 51Cr, 855r, or 465c; 3M Co., St Paul, MN) was randomized among the steps of the protocol. At the end of the experiment the pony was anesthetized with intravenous thiamylal sodium and killed by exsanguination. The brain, adrenal glands and kidneys were prepared for gamma nuclide spectroscopy as described previously (Manohar, 1986), The raw counts were corrected for background and crossover, and were compared with reference blood samples to determine blood flow (0) in ml. rain- 1. 100 g- 1 (Manohar, 1986; Parks and Manohar, 1983a,b). The calculation was performed as 0 = RR x CT/CR, where RR is the rate of reference sample withdrawal in mi. min- 1, and CT and CR respectively stand for radioactivity in the tissue and the reference blood. All criteria for tissue blood flow determination by the microsphere technique were completely satisfied (Buckberg et aL, 1971). The brain 02 supply was calculated as the product of regional brain blood flow and Cao~. Statisticalanalysis of the data. The data were subjected to two-way analysis of variance followed by Newman-Keuls multiple range test for comparison of treatment means (Steel and Torrie, 1960). A probability level of P < 0.05 was regarded as statistically significant. The data are presented as means + 1 SEM. Blood flow values for paired tissues (adrenal glands, kidneys, caudate nuclei, cerebral cortex, cerebral white matter) for each step of the protocol were compared using one-way analysis of variance, and significant differences were not found.

Results

In splenectomized ponies Pao2 remained near control value but Paco 2 and pH a decreased precipitously with both levels of exercise (table 1). The hemoglobin increased by 8 ~o and 13 % during sub-maximal and maximal exercise, respectively (table 1). The

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M. M A N O H A R TABLE 1

Some blood-gas and hemodynamic variables at rest and during exercise in nine healthy splenectomized ponies Rest (control)

Sub-maximal exercise

Maximal exercise

47 + 3

213 + 4**

222 + 4**

Mean arterial blood pressure (mm Hg)

118 + 6

141 + 5*

137 + 5*

Arterial pH

7.392 + 0.004

7.314 + 0.020*

7.227 + 0.013 **zx

Arterial O 2 tension(mmHg)

93

91

95

Arterial CO 2 tension (mm Hg)

39.4 + 0.6

30.9 + 1.3"*

29.4 + 0.9**

Hemoglobin concentration (g-d1-1)

12.3 + 0.4

13.3 + 0.5*

13.9 + 0.4*

Cardiac frequency (beats-min-1)

+3

+2

+2

* Significantly different from rest (control) at P < 0.05. ** Significantly different from rest (control) at P < 0.0001. z~ Significantly different from sub-maximal exercise at P < 0.005. TABLE 2 Blood flow ( m l - m i n - 1. 100 g - 1) in the paired tissues of nine healthy splenectomized ponies at rest and during exercise Tissues

Rest (control)

Sub-maximal exercise

Left cerebral cortex Right cerebral cortex

104.5 + 9.6 104.0 + 9.2

Maximal exercise

84.7 + 12.7 85.1 + 12.1

102.5 + 11.2 105.6 + 14.1

1.8 1.8

18.9 + 2.1 18.8 + 2.1

22.6 + 2.8 23.0 _+ 3.1

Left caudate nucleus Right caudate nucleus

125.6 + 10.2 126.2 + 12.5

100.9 + 11.2 98.3 + 12.8

119.2 + 17.4 132.0 + 18.2

Left adrenal gland Right adrenal gland

182.8 + 14.2 178.1 + 11.7

326.9 + 38.6* 304.5 + 40.4*

227.9 + 29.3 234.5 + 39.5

Left kidney Right kidney

1032 + 66 1028 + 60

346 + 110" 349 + 107"

Left cerebral white matter Right cerebral white matter

23.0 + 23.9 +

106 _+ 53 *zx 107 + 55 *zx

The kidneys were carbonized at 300 °C for 24 h before nuclide counting. All blood flow values are in m l . m i n - 1 . 100g -1. * Significantly different from rest (control) at P < 0.0001. zx Significantly different from sub-maximal exercise at P < 0.0001.

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81

,ooi El 41 so

60

•~=

w-

40

20

0

MEDULLA

PONS

160

~

MID-BRAIN

THAL-H,TH.

BRAIN-STEM

REST

¢

SUB-MAX. EX. MAX. EX.

140

~ = VS. REST (P<0.05)

120 100 8O

40 20 0

WHITE MATTER

]

CORTEX

CEREBELLUM

CAUDATE N.

[

CORTEX

WHITE MAI-rER

CEREBRUM

Fig. 1. Regional distribution of brain blood flow in nine healthy splenectomized ponies at rest (control), as well as during two levels of exercise (EX) performed on a treadmill. The treadmill speed used for maximal exercise (MAX EX) in the splenectomized ponies varied from 14 to 15 miles per hour. Asterisk denotes significant difference from control value. THAL-H.TH refers to thalamus-hypothalamus region of the brain. It should be noted that in the cerebellar cortex, blood flow was not different between sub-maximal (SUB-MAX) and maximal exercise.

Cao2 of splenectomized ponies, therefore, also increased from 16.1 + 0.6 ml- dl- 1 at rest to 17.3 + 0.6ml.d1-1 (by 7.5~) during sub-maximal exercise, and 18.0 + 0.6 ml.d1-1 (by 11.8~) with maximal exertion. The cardiac frequency and mean aortic pressure of splenectomized ponies increased to similar levels with both intensities of exercise (table 1). Whereas, the renal blood flow decreased during exercise, an increase in adrenal gland blood flow was observed during sub-maximal exercise (table 2). In the cerebral cortex, cerebral white matter and the caudate nuclei, blood flow (table 2) as well as the vascular resistance (table 3) remained similar between rest, sub-maximal and maximal exercise. In the cerebeUar cortex blood flow increased with exercise (39.6~ with sub-maximal and 51.9~o with maximal exercise; fig. 1) and the vascular resistance decreased from pre-exercise values (table 3). Significant changes were not observed in either the blood flow (fig. 1) or the vascular resistance (table 3) in the cerebellar white matter, medulla, pons, mid-brain and thalamus-hypothalamus with either intensity of exercise in splenectomized ponies. exercise (P < 0.05), but in all other regions of the brain 02 supply remained similar to the control (resting) values.

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M. MANOHAR TABLE 3

Vascular resistance (mm Hg/ml •rain- 1.g- 1) in various regions of the brain at rest and during exercise in nine splenectomized ponies Tissues

Rest (control)

Sub-maximal exercise

Maximal exercise

122 + 11 102 + 10 540 _+43

193 + 22 161 + 19 841 _+96

145 _+ 16 122 + 13 675 _+66

120 + 11 317 + 27

116_+ 12 410 + 50

92 + 8* 300 + 38

173 + 14 212 + 19 244 + 19

229 + 28 282 + 42 293 + 40

174 + 16 222 _+26 238 + 19

A. Cerebrum:

Cortex Caudate nuclei White matter B. Cerebellum:

Cortex White matter C. Brain-stem:

Thalamus-hypothalamus Pons Medulla

Vascular resistance was calculated as the quotient of mean aortic pressure (mm Hg) and tissue blood flow (ml. min - 1. g- l). * Significantly different from rest (control) at P < 0.05.

Discussion The b l o o d - g a s tensions and p H of exercising splenectomized ponies behaved quite similarly to those measured for exercising normal ponies (Parks and Manohar, 1983a,b; Manohar, 1986). The cardiac frequency elicited by maximal exertion in splenectomized ponies was not different from that for normal ponies (Manohar, 1986). It was also similar for the sub-maximal and maximal exercise steps (table 1). Persson and Lydin (1973) have reported that cardiac frequency response to sub-maximal exercise is exaggerated in splenectomized horses. The meager increase in hemoglobin concentration o f splenectomized ponies during strenuous exercise (table 1) is also consistent with previous reports (Persson and Lydin, 1973). (A) Regional cerebral circulation during exercise. The regional cerebral blood flow values in resting splenectomized ponies (table 2) were similar to those o f normal ponies (Manohar, 1986). Acute elevation o f blood pressure and hypocapnia are potent cerebral vasoconstrictive stimuli (Heistad and Kontos, 1983) that were evident in exercising splenectomized ponies (table 1). But regional cerebral blood flow (table 2) and 0 2 delivery remained unchanged from control values throughout the study. Since a tight coupling between cerebral O z supply and metabolic 0 2 d e m a n d is k n o w n to exist (Heistad and Kontos, 1983), it may be suggested that regional cerebral 0 2 requirements during exercise in splenectomized ponies were probably not different from the control

CEREBRAL CIRCULATIONDURING EXERCISE

83

values at rest. While the regional cerebral blood flow of normal ponies during exercise, in conformity with splenectomized ponies, remained similar to control (rest) values, the regional cerebral 0 2 delivery was increased as Cao2 rose by 5 8 ~ (Manohar, 1986). The results of the present study would imply that an augmentation of regional cerebral 0 2 delivery in normal exercising ponies (Manohar, 1986) may not necessarily reflect an augmentation of overall cerebral metabolism. The regional cerebral vascular resistance of splenectomized ponies (as in normal ponies; Manohar, 1986) also did not change with exercise (table 3). This happened despite the presence of marked hypocapnia and increased mean arterial pressure during exertion (table 1). Explanation for this may be related to the calculation of cerebral vascular resistance. The latter simply calculated as the quotient of mean aortic pressure and blood flow may not reflect true vascular resistance in cerebral vascular structures. Recently, the existence of a vascular waterfall (Starling's resistor) in the cerebral venous circulation has been suggested (Luce et al., 1982). (B) Cerebellar petfusion and 02 supply. As in normal ponies (Manohar, 1986), the exercise induced increments in cerebellar blood flow were limited to the gray matter in the splenectomized ponies (fig. 1). The cerebellar gray matter 0 2 delivery also increased (P < 0.05) during both intensities of exertion but that in the cerebellar white matter was not affected. We interpret this to mean that exercise in splenectomized ponies was attended by increased metabolic 0 2 demand only in the cerebeUar gray matter. In normal ponies, severe exercise had resulted in increased 0 2 supply to the cerebellar white matter as well because of a 58 ~ increase in Cao2. The data from splenectomized ponies would suggest that an augmentation of cerebellar white matter 02 supply in exercising normal ponies may also not necessarily reflect increased metabolic requirements. The cerebellar cortical blood flows at rest and during maximal exercise (105 + 8 and 160 + 18 ml. min - ~ • 100 g - ~; fig. 1) in splenectomized ponies were not different from the normal ponies (95 + 6 and 185 + 14 ml. min- 1. 100 g - 1 at rest and maximal exertion, respectively; Manohar, 1986). However, because of high Cao2 (22.4 + 0.4 vs 18.0 + 0.6 ml. dl- 1 in splenectomized ponies; P < 0.01), the cerebellar cortical 0 2 supply of normal ponies during maximal exercise exceeded that in splenectomized ponies (P < 0.01). Although maximal exercise in normal ponies was carried out at higher work intensity, it cannot be determined whether the entire increment in cerebellar cortical 0 2 supply of normal ponies during exercise was related to the augmented metabolism. (C) Regional brain-stem bloodflow and 02 supply. In normal ponies, blood flow and O2 supply increased in the medulla and pons during severe exertion (Manohar, 1986). This, however, did not happen in splenectomized ponies (fig. 1). The results in splenectomized ponies therefore, do not suggest an augmentation of metabolism in any portion of the brain-stem during exercise. In summary, these experiments demonstrated that with severe exercise in

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M. MANOHAR

splenectomized ponies, the blood flow and 02 supply increased only in the cerebeilar cortex. This means that metabolic 02 requirements of the cerebral and brain-stem structures during exercise probably did not differ from control. Thus, these data suggest that the increments in cerebral and brain-stem 02 supply in maximally exercising normal ponies (Manohar, 1986) may not necessarily reflect an increased metabolism in these regions.

Acknowledgements.The excellent technical assistance of Kelley Yorke, Tammy Baker, Dr. Jane Davis and Waiter Crackel is gratefully acknowledged. Thanks are due to the personnel at the Biomedical Communications Center for preparing the graph and at the Word Processing Center for typing the manuscript. This work was supported in part by grants-in-aid from the American Lung Association, the American Heart Association ( + Illinois Affiliate) and the Grayson Foundation.

References Bisgard, G.E., H.V. Forster, B. Byrnes, K. Stanek, J. Klein and M. Manohar (1978). Cerebrospinal fluid acid-base balance during muscular exercise. J. Appl. PhysioL 45: 94-101. Buckberg, G. D., J. C. Luck, D. B. Payne, J. I. E. Hoffman, J. P. Archie and D. E. Fixler (1971). Some sources of error in measuring regional blood flow with radioactive microspheres. J. Appl. Physiol. 31 : 598-604. Heistad, D.D. and H.A. Kontos (1983). Cerebral circulation. In: Handbook of Physiology. Section 2. The Cardiovascular System. Vol. Ili: Peripheral Circulation (Part I), edited by J.T. Shephard and F.M. Abboud. Washington, DC, American Physiological Society, pp. 137-182. Luce, J. M., J. S. Huseby, W. Kirk and J. Butler (1982). A starling resistor regulates cerebral venous outflow in dogs. J. Appl. PhysioL 53: 1496-1502. Manohar, M. (1986). Regional brain blood flow and 02 delivery during severe exertion in the pony. Respir. Physiol. 64: 339-349. Parks, C.M. and M. Manohar (1983a). Transmural coronary vasodilator reserve and flow distribution during severe exercise in ponies. J. Appl. Physiol. 54: 1641-1652; Parks, C. M. and M. Manohar (1983b). Distribution of blood flow during moderate and strenuous exercise in ponies (Equus caballus). Am. J. Vet. Res. 44: 1861-1866. Persson, S. G. B. and G. Lydin (1973). Circulatory effects of splenectomy in the horse. Zentr. Veterinaermed. Reihe A. 20: 521-530. Steel, R. G. D. and J.H. Torrie (1960). Principles and Procedures of Statistics. New York, McGraw-Hill, pp. 132-160.