Brain Research, 506 (1990) 149-152 Elsevier
149
BRES 23878
Changes in cerebrospinal fluid Na ÷ concentration do not underlie hypertensive responses to dietary NaCI in ,spontaneously hypertensive rats M.S. Mozaffari 1, S. Jirakulsomchok 1., S. Oparil 2, and J.M. Wyss ~'2 1Department of Cell Biology and Anatomy, and 2the Hypertension Research Program of the Department of Medicine, University of Alabama at Birmingham, Birmingham, AI 35294 (U.S.A.) (Accepted 12 September 1989) Key words: Sympathetic nervous system; Hypothalamus; Noradrenaline; Hypertension; Adrenoceptor
This study tests the hypothesis that dietary NaCI loading increases cerebrospinal fluid (CSF) Na ÷ concentration in NaCl-sensitive spontaneously hypertensive rats (SHR-S), resulting in an increase in arterial pressure. The high NaC1 diet caused a significant rise in systolic arterial pressure in SHR-S but not in normotensive Wistar Kyoto (WKY) rats. In contrast, the high NaCI diet caused a transient rise in CSF Na + that was similar in amplitude in SHR-S and WKY. A second experiment demonstrated that in SHR-S, concomitant dietary Ca2+ supplementation attenuated the dietary NaCl-induced exacerbation of hypertension, but did not alter the transient increase in CSF Na + concentration. Together, these results indicate that alterations in CSF Na ÷ concentration do not contribute to the increase in arterial pressure induced by a high NaC1 diet in SHR-S. Diets high in NaC1 exacerbate hypertension in NaC1sensitive spontaneously hypertensive rats (SHR-S) but do not elevate arterial pressure in Wistar-Kyoto rats (WKY) or NaCl-resistant S H R (SHR-R) 2°. Recent studies indicate that the nervous system plays an important role in the hypertensive response to supplemental dietary NaCI in SHR-S. Diets high in NaCI elevate sympathetic nervous system activity in SHR-S but not in W K Y or S H R - R 2a. In SHR-S but not in S H R - R or WKY, high NaC1 diets decrease noradrenaline turnover selectively in the anterior hypothalamic area ( A H A ) 3"22. The A H A contains neurons that have a sympathoinhibitory influence. Microinjection of a2 agonists into the A H A causes arterial pressure to decrease in normotensive and hypertensive rats 22, and electrical stimulation of this area results in decreases in both heart rate and arterial pressure 15, These observations are consistent with the hypothesis that reduced stimulation of central sympathoinhibitory neurons contributes to the rise in blood pressure in SHR-S on a high NaCI diet 3'21. Further evidence in support of this hypothesis comes from the findings that in SHR-S, dietary Ca 2÷ supplementation prevents both the decrease in noradrenaline turnover in A H A , and the increases in sympathetic nervous system activity and
blood pressure induced by dietary NaCI supplementation 21, None of these effects are observed in S H R - R or WKY. Thus, in the SHR-S on a high NaC1 diet, a decrease in A H A noradrenaline release and the resulting overactivity of the sympathetic nervous system appear to contribute to the elevation of blood pressure, but the mechanism(s) by which dietary NaC1 effects these changes remains unclear. The A H A lies close to the third ventricle and thus its neurons are exposed to changes in the ionic concentration of the cerebrospinal fluid (CSF). Small changes ( < 2 mEq) in CSF Na + concentration affect the firing rate of neurons in the paraventricular and supraoptic nuclei lz, and it seems likely that neurons in the nearby A H A may alter their firing rates following similar changes in CSF Na + concentration. Further, extracellular Na + excess diminishes the affinity of a2-adrenoceptors in vitro 7"s, and a similar effect in vivo could result in a reduced response of hypothalamic a2-adrenoceptors to noradrenaline, or a reduction in presynaptic inhibition mediated by the presynaptic a2 adrenoceptor. Gavras and his associates have suggested that these changes in central adrenoceptor affinity may contribute importantly to the hypertensive responses to dietary NaC1 in genetically predisposed individuals 5. The present study tests the
* Present address: Department of Biochemistry, Khon Kaen University, Khon Kaen, Thailand. Correspondence: J.M. Wyss, Department of Cell Biology & Anatomy, University of Alabama at Birmingham, University Station, Birmingham, AL 35294, U.S.A. 0006-8993/90/$03.50 © 1990 Elsevier Science Publishers B.V. (Biomedical Division)
150 hypothesis that diets high in NaCI exacerbate hypertension in SHR-S by increasing CSF Na + concentration and that the hypotensive effect of dietary Ca 2+ is mediated by a reduction in CSF Na + concentration. SHR-S (IBU-3 colony) and WKY were obtained from Taconic Farms (Germantown, NY) at 7 weeks of age 21. All rats were maintained 5 per cage at constant humidity (60 + 5%), temperature (24 + 1 °C) and light cycle (06.00-18.00 h). Three days after arrival, systolic arterial pressure was measured in conscious, prewarmed rats by tail plethysmography, following which the rats were randomly assigned to special diet groups. In Expt. 1, groups (n = 10) of SHR-S and WKY were placed on a basal NaCI (0.75% NaCI, 0.68% Ca 2÷) or a high NaCI (8% NaC1, 0.68% Ca 2÷) diet (Teklad Madison, WI). In the second study, SHR-S were placed on a high Ca z+ (0.75% NaCI, 2% Ca 2+) or high NaC1 and high Ca 2÷ (8% NaC1, 2% Ca 2÷) diet. These diets have been described in detail elsewhere 21. One, 3, 7 and 14 days after the initiation of the diets, CSF samples were withdrawn from the cisterna magna by the method described by Frankmann 4. Briefly, rats were anesthetized with sodium pentobarbital (40 mg/kg body weight; i.p.) and their heads were mounted in a stereotaxic frame. A small, midsagittal incision was made through the skin approximately 7 mm below the occipital crest, and a 28-gauge needle was advanced into the cisterna magna with a micromanipulator. After the needle entered the cisterna magna, CSF was allowed to flow freely (aided by gravity) through the needle and attached tubing until approximately 150 #1 were collected in a microvial (typically 15 min). The needle then was withdrawn and the skin incision sutured. The success rate for obtaining CSF samples that were not contaminated ,with blood was greater than 95%. CSF samples which contained a trace of blood were eliminated from the subsequent analysis. One, 3, 7 and 14 days after initiation of the diets, approximately 0.4 ml of blood was drawn from the tail vein for analysis of serum Na + concentration, and a small sample was obtained in heparinized capillary tubes for determination of hematocrit. Na + and K ÷ concentrations in CSF and serum were determined by flame photometry. Systolic arterial pressure was measured 2 weeks after initiation of dietary regimens by the tail cuff method. All data were analyzed by multivariant analysis of variance with appropriate post-hoc tests to determine the source of significant main effects or interactions. Prior to initiation of the experimental diets, SHR-S were significantly hypertensive (systolic arterial pressure [SAP] = 156 _ 3 mm Hg). In SHR-S on the basal NaCI diet, arterial pressure continued to increase during the next 2 weeks, but the high (compared to basal) NaCI diet caused a much greater increase in arterial pressure (SAP
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L2] WKY 1% NoCI WKY 8% NoCI ~:3 SHR-S 1% NoC~ E'23 SHR-S 8% NoCI SHR-S I% NoCI Z 2,o~Ca +÷ [Z_q SHR-S 8% NoCl/ 2% Ca ++
149
147
O
145
3
7
t4
Time ( d a y s )
Fig. 1. Cerebrospinai fluid sodium concentration of SHR-S and WKY 1, 3, 7, and 14 days after initiation of the special diets. *P < 0.05 compared to 1% NaCl diet group of the same strain and on the same Ca2+ diet.
= 195 _+ 4 mm Hg; 173 +_ 3 m m Hg, respectively). The high NaCI diet had no effect on arterial pressure in WKY (final SAP = basal NaCl diet 114 + 4 mm ~ ; : h i g h NaCl diet 117 _+ 3 m m Hg). In SHR-S on the high (compared to basal) NaCl diet, CSF Na + concentration was elevated on day 1, but not thereafter (Fig. 1). In WKY, the high (compared to basal) NaCl diet caused a similar transient increase in CSF Na + concentration (Fig. 1). There was no significant difference between SHR-S and WKY in these responses. CSF K + concentrations were not altered by dietary NaCI. One and two weeks exposure to the high (compared to basal) NaCl diet increased plasma Na ÷ concentration in SHR-S but not in WKY (Fig. 2). In SHR-S the high NaC1 and Ca 2+ (compared to basal NaCl) diet did not increase SAP (final SAP -- 173 _+ 2 mm Hg), but did increase CSF Na + to the same extent as the high NaCl diet. In SHR-S on the high Ca 2+ diet, arterial pressure rose significantly less (final SAP = 163
~--
ESZ] WKY 1% NaCI [ ~ WKY 8~ NaCI FL~7 SHR-S 1% NoCt SHR-S 8% NoCl ~ SHR-S 1Z NaCIZ 2% Ca++ I--'2"1 SHR-S 87** NaCI/ 2% Co++
150
E *
2
HRJ
o o + Z
(o
130
1
3 Time
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14
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Fig. 2. Plasma sodium concentration in SHR-S and WKY 1, 3, 7, and 14 days after initiation of the special diets. *P < 0.05 ~mpared to 1% NaCl diet group of the same strain and on the same Ca2+ diet.
151 +- 3) than in the other groups of SHR-S. SHR-S on the high NaCI and Ca 2÷ (compared to the high Ca 2÷) diet displayed transient elevation of plasma Na + concentration. Hematocrit was not altered significantly by any of the diets in SHR-S or WKY. Several lines of evidence suggest that increased CSF Na ÷ concentration contribute to dietary NaCl-induced increases in blood pressure. First, an elevation in CSF Na ÷ concentration plays a pivotal role in the initiation of several models of low renin hypertension in rats including deoxycorticosterone acetate-NaC1 (DOCA-NaCI) and one-kidney Goldblatt hypertension l°'n. Second, a significant rise in CSF Na + concentration and osmolality occurs 3 days after the initiation of one-kidney, one-wrap Grollman renal hypertension, and this may be a primary stimulus for the elevation of arterial pressure in that model 9. Third, a number of studies carried out in normotensive rats demonstrated that intracerebroventricular administration of hypertonic saline causes a hypertensive response that is attributable to a centrally mediated increase in peripheral sympathetic nervous system activity 2'6A3-15. The mechanism(s) of the pressor response to central administration of hypertonic NaCI is not understood fully, but several studies suggest that the principal site of action of hypertonic saline is within the anterior hypothalamus. First, intracerebroventricular injection of hypertonic saline reduces sympathoinhibitory responses elicited by electrical stimulation of the A H A 15. Second, the A H A lies near the third ventricle and thus is likely affected by changes in CSF Na ÷ concentration, and third, neurons in this region are known to play an important role in the central regulation of cardiovascular and renal function ~5. In contrast, a recent study by Takada et al. 19 suggests that increases in CSF Na ÷ do not contribute to deoxycorticosterone acetate-NaC1 (DOCA-NaCI) hypertension, since there is no relationship between systolic arterial pressure and CSF Na ÷ concentration at any stage in the development of DOCA-NaC1 hypertension. I n Dahl NaCl-sensitive rats on a high compared to basal NaCI diet, CSF Na ÷ concentration is elevated, but only after the NaC1 has increased arterial pressure for several days, indicating that the increase in CSF Na ÷ does not trigger hypertension in this model 16. The current results indicate that changes in CSF Na ÷ concentration do not mediate NaCl-exacerbated hypertension in SHR-S. Although a transient increase in CSF Na ÷ concentration was noted in SHR-S on the high NaCI diet, WKY rats on the high NaCI diet displayed an equivalent rise in CSF Na + concentration. In contrast, the high NaCI diet exacerbated hypertension in SHR-S but had no effect on arterial pressure in normotensi-
ve WKY rats. Concomitant dietary Ca 2÷ supplementation prevented the full expression of the hypertensive effects of the high dietary NaC1 in SHR-S, but did not alter the transient elevation of CSF Na +. Finally, in SHR-S on a high NaCl diet, the transient rise in CSF Na ÷ occurred at least 4 days prior to the NaCl-induced rise in arterial pressure 3'21. Dietary NaCl caused a progressive elevation of plasma Na ÷ concentration in SHR-S but not in WKY. The rise in plasma Na ÷ concentration was significant at day 7, a time at which CSF Na + concentration had returned to normal in SHR-S and WKY on the high NaC1 diet. The lack of an increase in plasma Na ÷ level at a time when CSF Na ÷ concentration was elevated indicates that the increases in CSF Na + concentration are independent of changes in plasma Na + concentration. Taken together, our data do not support the hypothesis that alterations in CSF Na ÷ concentration mediate the dietary NaClinduced changes in hypothalamic noradrenaline turnover, sympathetic nervous system activity or arterial pressure in SHR-S 3'21. Expansion of CSF volume, irrespective of CSF Na ÷ concentration, may play a role in pathogenesis of hypertension in SHR-S on a high NaCI diet. Ritter and colleagues demonstrated that the brain ventricles are significantly larger in SHR than WKY and that the circumventricular regions of the brain are significantly smaller in SHR than WKY 17'18. Lowering arterial pressure of SHR does not decrease the cerebroventricular expansion in SHR TM, and raising arterial pressure in SHR-S by administering a high NaC1 diet for 2 weeks does not increase ventricular size significantly (unpublished results from our laboratory). More accurate methods of measuring CSF volume in the rat are needed to assess the importance of these changes. In summary, transient elevations in CSF Na + concentration accompany dietary NaCI loading of both SHR-S and WKY, but the high NaCI diet exacerbates hypertension only in SHR-S. In SHR-S, simultaneous dietary Ca 2÷ supplementation attenuates the dietary NaCl-induced rise in arterial pressure but does not reduce the transient increase in CSF Na ÷. The lack of a correlation between changes in CSF Na + concentration and arterial pressure in SHR-S on the special diets strongly suggests that" alterations in CSF Na + concentration do not contribute to the NaCl-induced exacerbation of hypertension in SHR-S.
This work was supported in part by National Heart, Lung and Blood Institutes Grants HL 37722, HL 22544, HL 25452, HL 36390, HL 35051, and by a grant from the National Dairy Board and administered in cooperation with National Dairy Council.
152 1 Buggy, J. and Johnson, A.K., Preoptic-hypothalamic periventricular lesions: thirst deficits and hypernatremia, Am. J. Physiol., 233 (1977) R44-R52. 2 Bufiag, R.D. and Miyajima, E., Sympathetic hyperactivity elevates blood pressure during acute cerebroventricular infusions of hypertonic salt in rats, J. Cardiovas. Pharmacol., 6 (1984) 844-851. 3 Chen, Y.E, Meng, Q., Wyss, J.M., Jin, H. and Oparil, S., High NaCI diet reduces hypothalamic norepinephrine turnover in hypertensive rats, Hypertension, 24 11 (1988) 55-62. 4 Frankmann, S.E, A technique for repeated sampling of CSF from the anesthetized rat, Physiol. Behav., 37 (1986) 489-493. 5 Gavras, H., How does salt raise blood pressure? Hypertension, 8 (1986) 83-88. 6 Gavras, H., Bain, G.T., Bland, L.I., Vlahakos, D. and Gavras, I., Hypertonic response to saline microinjection in the area of the nucleus tractus solitarii of the rat, Brain Research, 343 (1985) 113-119. 7 Glossman, H., Lubbecke, E, Beltmann, P., Sattler, E.L. and Doell, G., Ionic modulation of alpha-adrenoceptors, J. Cardiovas. Pharrnacol., 4 (1982) 551-557. 8 Greenberg, D.A., U'prichard, D.C., Sheehan, EO. and Snyder, S.H., Alpha-adrenergic receptors in the brain: differential effects of sodium ion on binding of [3H]-agonists and [3HIantagonists, Brain Research, 140 (1978) 378-384. 9 Haywood, J.R., Buggy, J., Fink, G.D., DiBona, G.E, Johnson, A.K. and Brody, M.J., Alterations in cerebrospinal fluid sodium and osmolality in rats during one-kidney, one-wrap renal hypertension, Clin. Exp. Pharmaeol. Physiol., 11 (1984) 545549. 10 Haddy, F.J., Pamnani, M.B. and Clough, D.L., Humoral factors and the sodium-potassium pump in volume expanded hypertension, Life Sci., 24 (1986) 2105-2118. 11 Jandhyala, B.S., Is expansion of extracellular fluid volume essential for the development of 'volume-expanded' hypertension? Clin. Exp. Theory Practice, A8 (1986) 457-472. 12 Joynt, R.J., Verney's concept of the osmoreeeptor, Arch. Neurol., 14 (1966) 331-344.
13 Kawano, Y. and Ferrario, M., Neurohormonal characteristics oi cardiovascular response to intraventricular hypertonic NaCI, Am. J. Physiol., 247 (1984) H422-H428. 14 Kohlmann Jr., O., Gavras, I., BioUaz, J., Biollaz, B. and Gavras, H., Sodium chloride-induced partial inhibition in vivo of alphaz-adrenoceptor agonist function, J. Hypertension, 3 (1985) 269-274. 15 Miyajima, E. and Bunag, R.D., Chronic cerebroventricular infusion of hypertonic sodium chloride in rats reduces hypothalamic sympatho-inhibition and elevates blood pressure, Circ. Res., 54 (1984) 556-575. 16 Nakamura, K. and Cowley, A.W., Sequential changes of cerebrospinal fluid sodium during the development of hypertension in Dahl rats, Hypertension, 13 (1989) 243-249. 17 Ritter, S. and Dinh, T.T., Progressive postnatal dilation of brain ventricles in spontaneously hypertensive rats, Brain Research. 370 (1986) 327-332. 18 Ritter, S., Dinh, T.T., Stone, S. and Ross, N., Cerebroventricular dilation in spontaneously hypertensive rats (SHR-S) is not attenuated by reduction of blood pressure, Brain Research, 450 (1988) 354-359. 19 Takata, Y., Yamashita, Y., Takishita, S. and Fujishima, M., Lack of increase in concentrations of cerebrospinal fluid sodium in rats with various stages of DOCA-salt hypertension, Life Sci., 42 (1982) 1223-1229. 20 Wyss, J.M., Chen, Y.E, Jin, H., Gist, R. and Oparil, S., Spontaneously hypertensive rats exhibit reduced hypothalamic noradrenergic input after NaCI loading, Hypertension, 10 (1987) 313-320. 21 Wyss, J.M., Chen, Y.-E, Meng, M., Jin, H., Jirakulsomchok, S. and Oparil, S., Dietary Ca2+ prevents NaCl-induced exacerbation of hypertension and increases hypothalamic NE turnover in SHR-S, J. Hypertension, in press. 22 Wyss, J.M., Yang, R., Jin, H. and Oparil, S., Hypothatamic microinjection of aipha2-adrenoceptor agonists causes greater sympathoinhibition in spontaneously hypertensive rats on high NaCI diets, J. Hypertension, 6 (1988) 805-813.
149
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Changes in cerebrospinal fluid Na ÷ concentration do not underlie hypertensive responses to dietary NaCI in ,spontaneously hypertensive rats M.S. Mozaffari 1, S. Jirakulsomchok 1., S. Oparil 2, and J.M. Wyss ~'2 1Department of Cell Biology and Anatomy, and 2the Hypertension Research Program of the Department of Medicine, University of Alabama at Birmingham, Birmingham, AI 35294 (U.S.A.) (Accepted 12 September 1989) Key words: Sympathetic nervous system; Hypothalamus; Noradrenaline; Hypertension; Adrenoceptor
This study tests the hypothesis that dietary NaCI loading increases cerebrospinal fluid (CSF) Na ÷ concentration in NaCl-sensitive spontaneously hypertensive rats (SHR-S), resulting in an increase in arterial pressure. The high NaC1 diet caused a significant rise in systolic arterial pressure in SHR-S but not in normotensive Wistar Kyoto (WKY) rats. In contrast, the high NaCI diet caused a transient rise in CSF Na + that was similar in amplitude in SHR-S and WKY. A second experiment demonstrated that in SHR-S, concomitant dietary Ca2+ supplementation attenuated the dietary NaCl-induced exacerbation of hypertension, but did not alter the transient increase in CSF Na + concentration. Together, these results indicate that alterations in CSF Na ÷ concentration do not contribute to the increase in arterial pressure induced by a high NaC1 diet in SHR-S. Diets high in NaC1 exacerbate hypertension in NaC1sensitive spontaneously hypertensive rats (SHR-S) but do not elevate arterial pressure in Wistar-Kyoto rats (WKY) or NaCl-resistant S H R (SHR-R) 2°. Recent studies indicate that the nervous system plays an important role in the hypertensive response to supplemental dietary NaCI in SHR-S. Diets high in NaCI elevate sympathetic nervous system activity in SHR-S but not in W K Y or S H R - R 2a. In SHR-S but not in S H R - R or WKY, high NaC1 diets decrease noradrenaline turnover selectively in the anterior hypothalamic area ( A H A ) 3"22. The A H A contains neurons that have a sympathoinhibitory influence. Microinjection of a2 agonists into the A H A causes arterial pressure to decrease in normotensive and hypertensive rats 22, and electrical stimulation of this area results in decreases in both heart rate and arterial pressure 15, These observations are consistent with the hypothesis that reduced stimulation of central sympathoinhibitory neurons contributes to the rise in blood pressure in SHR-S on a high NaCI diet 3'21. Further evidence in support of this hypothesis comes from the findings that in SHR-S, dietary Ca 2÷ supplementation prevents both the decrease in noradrenaline turnover in A H A , and the increases in sympathetic nervous system activity and
blood pressure induced by dietary NaCI supplementation 21, None of these effects are observed in S H R - R or WKY. Thus, in the SHR-S on a high NaC1 diet, a decrease in A H A noradrenaline release and the resulting overactivity of the sympathetic nervous system appear to contribute to the elevation of blood pressure, but the mechanism(s) by which dietary NaC1 effects these changes remains unclear. The A H A lies close to the third ventricle and thus its neurons are exposed to changes in the ionic concentration of the cerebrospinal fluid (CSF). Small changes ( < 2 mEq) in CSF Na + concentration affect the firing rate of neurons in the paraventricular and supraoptic nuclei lz, and it seems likely that neurons in the nearby A H A may alter their firing rates following similar changes in CSF Na + concentration. Further, extracellular Na + excess diminishes the affinity of a2-adrenoceptors in vitro 7"s, and a similar effect in vivo could result in a reduced response of hypothalamic a2-adrenoceptors to noradrenaline, or a reduction in presynaptic inhibition mediated by the presynaptic a2 adrenoceptor. Gavras and his associates have suggested that these changes in central adrenoceptor affinity may contribute importantly to the hypertensive responses to dietary NaC1 in genetically predisposed individuals 5. The present study tests the
* Present address: Department of Biochemistry, Khon Kaen University, Khon Kaen, Thailand. Correspondence: J.M. Wyss, Department of Cell Biology & Anatomy, University of Alabama at Birmingham, University Station, Birmingham, AL 35294, U.S.A. 0006-8993/90/$03.50 © 1990 Elsevier Science Publishers B.V. (Biomedical Division)
150 hypothesis that diets high in NaCI exacerbate hypertension in SHR-S by increasing CSF Na + concentration and that the hypotensive effect of dietary Ca 2+ is mediated by a reduction in CSF Na + concentration. SHR-S (IBU-3 colony) and WKY were obtained from Taconic Farms (Germantown, NY) at 7 weeks of age 21. All rats were maintained 5 per cage at constant humidity (60 + 5%), temperature (24 + 1 °C) and light cycle (06.00-18.00 h). Three days after arrival, systolic arterial pressure was measured in conscious, prewarmed rats by tail plethysmography, following which the rats were randomly assigned to special diet groups. In Expt. 1, groups (n = 10) of SHR-S and WKY were placed on a basal NaCI (0.75% NaCI, 0.68% Ca 2÷) or a high NaCI (8% NaC1, 0.68% Ca 2÷) diet (Teklad Madison, WI). In the second study, SHR-S were placed on a high Ca z+ (0.75% NaCI, 2% Ca 2+) or high NaC1 and high Ca 2÷ (8% NaC1, 2% Ca 2÷) diet. These diets have been described in detail elsewhere 21. One, 3, 7 and 14 days after the initiation of the diets, CSF samples were withdrawn from the cisterna magna by the method described by Frankmann 4. Briefly, rats were anesthetized with sodium pentobarbital (40 mg/kg body weight; i.p.) and their heads were mounted in a stereotaxic frame. A small, midsagittal incision was made through the skin approximately 7 mm below the occipital crest, and a 28-gauge needle was advanced into the cisterna magna with a micromanipulator. After the needle entered the cisterna magna, CSF was allowed to flow freely (aided by gravity) through the needle and attached tubing until approximately 150 #1 were collected in a microvial (typically 15 min). The needle then was withdrawn and the skin incision sutured. The success rate for obtaining CSF samples that were not contaminated ,with blood was greater than 95%. CSF samples which contained a trace of blood were eliminated from the subsequent analysis. One, 3, 7 and 14 days after initiation of the diets, approximately 0.4 ml of blood was drawn from the tail vein for analysis of serum Na + concentration, and a small sample was obtained in heparinized capillary tubes for determination of hematocrit. Na + and K ÷ concentrations in CSF and serum were determined by flame photometry. Systolic arterial pressure was measured 2 weeks after initiation of dietary regimens by the tail cuff method. All data were analyzed by multivariant analysis of variance with appropriate post-hoc tests to determine the source of significant main effects or interactions. Prior to initiation of the experimental diets, SHR-S were significantly hypertensive (systolic arterial pressure [SAP] = 156 _ 3 mm Hg). In SHR-S on the basal NaCI diet, arterial pressure continued to increase during the next 2 weeks, but the high (compared to basal) NaCI diet caused a much greater increase in arterial pressure (SAP
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149
147
O
145
3
7
t4
Time ( d a y s )
Fig. 1. Cerebrospinai fluid sodium concentration of SHR-S and WKY 1, 3, 7, and 14 days after initiation of the special diets. *P < 0.05 compared to 1% NaCl diet group of the same strain and on the same Ca2+ diet.
= 195 _+ 4 mm Hg; 173 +_ 3 m m Hg, respectively). The high NaCI diet had no effect on arterial pressure in WKY (final SAP = basal NaCl diet 114 + 4 mm ~ ; : h i g h NaCl diet 117 _+ 3 m m Hg). In SHR-S on the high (compared to basal) NaCl diet, CSF Na + concentration was elevated on day 1, but not thereafter (Fig. 1). In WKY, the high (compared to basal) NaCl diet caused a similar transient increase in CSF Na + concentration (Fig. 1). There was no significant difference between SHR-S and WKY in these responses. CSF K + concentrations were not altered by dietary NaCI. One and two weeks exposure to the high (compared to basal) NaCl diet increased plasma Na ÷ concentration in SHR-S but not in WKY (Fig. 2). In SHR-S the high NaC1 and Ca 2+ (compared to basal NaCl) diet did not increase SAP (final SAP -- 173 _+ 2 mm Hg), but did increase CSF Na + to the same extent as the high NaCl diet. In SHR-S on the high Ca 2+ diet, arterial pressure rose significantly less (final SAP = 163
~--
ESZ] WKY 1% NaCI [ ~ WKY 8~ NaCI FL~7 SHR-S 1% NoCt SHR-S 8% NoCl ~ SHR-S 1Z NaCIZ 2% Ca++ I--'2"1 SHR-S 87** NaCI/ 2% Co++
150
E *
2
HRJ
o o + Z
(o
130
1
3 Time
7
14
(days)
Fig. 2. Plasma sodium concentration in SHR-S and WKY 1, 3, 7, and 14 days after initiation of the special diets. *P < 0.05 ~mpared to 1% NaCl diet group of the same strain and on the same Ca2+ diet.
151 +- 3) than in the other groups of SHR-S. SHR-S on the high NaCI and Ca 2÷ (compared to the high Ca 2÷) diet displayed transient elevation of plasma Na + concentration. Hematocrit was not altered significantly by any of the diets in SHR-S or WKY. Several lines of evidence suggest that increased CSF Na ÷ concentration contribute to dietary NaCl-induced increases in blood pressure. First, an elevation in CSF Na ÷ concentration plays a pivotal role in the initiation of several models of low renin hypertension in rats including deoxycorticosterone acetate-NaC1 (DOCA-NaCI) and one-kidney Goldblatt hypertension l°'n. Second, a significant rise in CSF Na + concentration and osmolality occurs 3 days after the initiation of one-kidney, one-wrap Grollman renal hypertension, and this may be a primary stimulus for the elevation of arterial pressure in that model 9. Third, a number of studies carried out in normotensive rats demonstrated that intracerebroventricular administration of hypertonic saline causes a hypertensive response that is attributable to a centrally mediated increase in peripheral sympathetic nervous system activity 2'6A3-15. The mechanism(s) of the pressor response to central administration of hypertonic NaCI is not understood fully, but several studies suggest that the principal site of action of hypertonic saline is within the anterior hypothalamus. First, intracerebroventricular injection of hypertonic saline reduces sympathoinhibitory responses elicited by electrical stimulation of the A H A 15. Second, the A H A lies near the third ventricle and thus is likely affected by changes in CSF Na ÷ concentration, and third, neurons in this region are known to play an important role in the central regulation of cardiovascular and renal function ~5. In contrast, a recent study by Takada et al. 19 suggests that increases in CSF Na ÷ do not contribute to deoxycorticosterone acetate-NaC1 (DOCA-NaCI) hypertension, since there is no relationship between systolic arterial pressure and CSF Na ÷ concentration at any stage in the development of DOCA-NaC1 hypertension. I n Dahl NaCl-sensitive rats on a high compared to basal NaCI diet, CSF Na ÷ concentration is elevated, but only after the NaC1 has increased arterial pressure for several days, indicating that the increase in CSF Na ÷ does not trigger hypertension in this model 16. The current results indicate that changes in CSF Na ÷ concentration do not mediate NaCl-exacerbated hypertension in SHR-S. Although a transient increase in CSF Na ÷ concentration was noted in SHR-S on the high NaCI diet, WKY rats on the high NaCI diet displayed an equivalent rise in CSF Na + concentration. In contrast, the high NaCI diet exacerbated hypertension in SHR-S but had no effect on arterial pressure in normotensi-
ve WKY rats. Concomitant dietary Ca 2÷ supplementation prevented the full expression of the hypertensive effects of the high dietary NaC1 in SHR-S, but did not alter the transient elevation of CSF Na +. Finally, in SHR-S on a high NaCl diet, the transient rise in CSF Na ÷ occurred at least 4 days prior to the NaCl-induced rise in arterial pressure 3'21. Dietary NaCl caused a progressive elevation of plasma Na ÷ concentration in SHR-S but not in WKY. The rise in plasma Na ÷ concentration was significant at day 7, a time at which CSF Na + concentration had returned to normal in SHR-S and WKY on the high NaC1 diet. The lack of an increase in plasma Na ÷ level at a time when CSF Na ÷ concentration was elevated indicates that the increases in CSF Na + concentration are independent of changes in plasma Na + concentration. Taken together, our data do not support the hypothesis that alterations in CSF Na ÷ concentration mediate the dietary NaClinduced changes in hypothalamic noradrenaline turnover, sympathetic nervous system activity or arterial pressure in SHR-S 3'21. Expansion of CSF volume, irrespective of CSF Na ÷ concentration, may play a role in pathogenesis of hypertension in SHR-S on a high NaCI diet. Ritter and colleagues demonstrated that the brain ventricles are significantly larger in SHR than WKY and that the circumventricular regions of the brain are significantly smaller in SHR than WKY 17'18. Lowering arterial pressure of SHR does not decrease the cerebroventricular expansion in SHR TM, and raising arterial pressure in SHR-S by administering a high NaC1 diet for 2 weeks does not increase ventricular size significantly (unpublished results from our laboratory). More accurate methods of measuring CSF volume in the rat are needed to assess the importance of these changes. In summary, transient elevations in CSF Na + concentration accompany dietary NaCI loading of both SHR-S and WKY, but the high NaCI diet exacerbates hypertension only in SHR-S. In SHR-S, simultaneous dietary Ca 2÷ supplementation attenuates the dietary NaCl-induced rise in arterial pressure but does not reduce the transient increase in CSF Na ÷. The lack of a correlation between changes in CSF Na + concentration and arterial pressure in SHR-S on the special diets strongly suggests that" alterations in CSF Na + concentration do not contribute to the NaCl-induced exacerbation of hypertension in SHR-S.
This work was supported in part by National Heart, Lung and Blood Institutes Grants HL 37722, HL 22544, HL 25452, HL 36390, HL 35051, and by a grant from the National Dairy Board and administered in cooperation with National Dairy Council.
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