Acta Physiol Scand 1992, 145, 297-298
Is autoregulation of cerebral blood flow in rats influenced by nitro-L-arginine, a blocker of the synthesis of nitric oxide? Q WANG, "0.B. P A U L S O N and N. A. L A S S E N Department of Clinical Physiology and Nuclear Medicine, Bispebjerg Hospital, "Department of Neurology, Rigshospital, Denmark Autoregulation of blood flow is the term used to describe the constancy of organ blood flow observed when the arterial blood pressure is varied. It implies constriction of the resistance vessels, the arterioles, when the pressure increases and their dilatation when the pressure decreases. Autoregulation is seen in all organs of the body, but it has been studied in the cerebral circulation in particular (Lassen 1964, Paulson et al. 1990, Schmidt et al. 1990). Many theories have been proposed to explain the response. Most authors have favoured the myogenic theory. It holds that the smooth-muscle cells of the arterial wall are alone in eliciting the response. These cells are assumed to 'sense' the increased stretch caused by elevation of the blood pressure and to respond to this stretch by a constriction and vice versa. Recently, however, evidence has been published pointing to a key role of the arterial endothelium in autoregulation of CBF and probably also of blood flow through other organs. Studies on isolated segments of arteries from the brain of cats and dogs have shown that the normal increase in vascular tone to stretch is abolished if the endothelium is removed (Harder 1987, Katusic et al. 1987). In studies on the diameter of the pial arterioles of cats it was demonstrated that damage to the endothelium caused by ultraviolet light abolished the normal vasodilator response to a reduced blood pressure, while the same vessels have a preserved vasodilatation to hypercapnia (Gotoh et al. 1987). A number of vasoactive substances are now known to be produced by the endothelium. They comprise both potent vasoconstrictors and potent vasodilators. In particular the very rapidly cleared endotheliumderived relaxing factor (nitric oxide) (Palmer et al. 1987) might seem to be a suitable candidate, as it would explain the short time constant of the autoregulatory response. It is of interest, therefore, to Received 6 March 1992, accepted 30 March 1992. Key words : autoregulation, cerebral blood flow (CBF), nitric oxide (NO), nitro-1-arginine (NOLAG). Correspondence : Niels A. Lassen, Department of Clinical Physiology and Nuclear Medicine, Bispebjerg Hospital, DK-2400, Copenhagen NV, Denmark.
study cerebral autoregulation when nitric oxide (NO) synthesis is blocked. Nitro-L-arginine (NOLAG), a potent inhibitor of N O synthesis, was used in the present study. Its effect on CBF autoregulation was studied in anaesthetized (0.6% halothane/70% N,O), artificially ventilated Wistar rats. Cerebral blood flow (CBF) was measured by the intra-carotid '33Xe injection method as adapted for small rodents (Hertz et al. 1977).
50
I00 MABP (mmHg)
I50
. . . . . . . . . . . . . . . . . . . . . 50 100 I50 MABP (mmHg)
Fig. 1. Cerebral blood flow autoregulation in (a) control (saline) and (b) NOLAG-treated animals. CBF is given as a percentage of baseline values (in NOLAG group, the first measured CBF after NOLAG served as baseline). The autoregulation curve was drawn and low limits were calculated by a computerized procedure (see text).
297
298
Q. Mnng, 0. B . Paulson und R;. A. Lassen
The loner limit of CBF autoregulation \\as studied in 17 rats. Following measurement of baseline CBF, nine rats \\ere given 7.5 mg kg ' SOL.-\G. 0.1 ml min for h min intracarotidly. The remaining eight served as control, infusing only the vehicle (saline). In the control animals \I.IRP was raised gradual]!- to a level 20- 30 n m H g abo\-e baseline \l.IRP b! an infusion of noradrenaline (?---to x IO-" nig min I ) in order to study the expected plateau of CBF autoregulation. The blood pressure n a s then allowed to drop slowly by controlled bleeding t o the lovest measurdhk h e ! . CBF u a s measured a t approsimately 10 mmHg intervals of JLIBP during the whole stud!. In NOI.r\G-treated animals, the increase in blood pressure a a s noted. rendering it unnecessary to use noradrenaline. The autoregulation curl-e \\as calculated from the pooled CBF and .\l.iBP data in each group of animals, using a computerized procedure described else\\ here (Schmidt et ti(. 1990). Throughout the M hole experiment, bod! temperature, arterial PaO,, PaCO?, p€I were continuously monitored and maintained in the physiological range. There I\ ere no signiticant differences bet\\een saline and SOT. I G groups in this respect. The results showed that in all rats autoregulation of CBF \\-as preser\-ed (Fig. 1). NOI..\G (7.5 mg kg ') elicited an increase in \I.IBP h! .3h",, and a decrease in CBF b! 23",, as u e had seen in a previous stud!(\\'ang t t u/. 1991). .It this lon-ered CBF l e d the studied autoregulation of CBF was intact, as practicall! the same lover limit was observed in both groups, i 6 & 3.4 mmfIg for saline- and 78 6.1 mndlg for SOI,.-\G-treated animals (mean iSE, P > 0.05). In a pre\-ious stud!- we have shown, that the same dose i d N 0 1 , I G of 7..i mg kg \\-ill cause a marked suppression of the carbon dioside responsiveness of CHF (\\rang t! a/. 1991). The effects on SLIBP and CBF seen in this stud!- also constitute el-idence of the biological dctit-ity of the dose gi\en, effects that pe for at least 2 h. On this basis we conclude that 5: nthase actii-it! \\as indeed eKecti\-el!- inhibited in our studies. The tindings of a preseried CBF autoregulation consequentl! mean that this regulation is probahl! not dependent on endothelium-deriwd nitric oxide. Ltl'hen 30 mg kg ' NOLriG \\-as infused in our earlier stud! (M'ang el L i / . 1991), ~e observed an increase in blood pressure \$ hich exceeded the upper limit of(:HF autoregulation ( > 150 mmIIg) (Paulson r t nl. 1990). In some rats the blood pressure rose as high as 170 mmfIg, but CBF still remained constant (unpublished observation). Thus the autoregulator! plateau seems widened in the high-pressure range, as it does in the case of low CBF caused by hyperLentilation. .Is noted in the introduction, remoial of the
endothelium abolishes the normal constrictor response of isolated cerehral \-essels to stretch (Harder 1987, Katusic el izl. I%;), and destruction of the endothelium in a pial vessel abolishes the normal dilation response resulting from a drop in blood pressure (Gotoh et ctl. 1987). Our demonstration of preserved CBF autoregulation despite NO synthase blockage suggests, therefore, that the results mentioned ma! he due to elimination o f other endothelium-deriwd factors, rather than EDRF (NO). The stud?- was supported bl; the Alfred Benzon Foundation, the Lundbeck Foundation and the Danish Health Xledical Council, Denmark. The escellent technical assistance of Kim Pape Gadegdard and Pia Tejmer is gratefully acknowledged.
REFERENCES F., F L K W C I I I , y.,h I A N O , T., TANAK.4, K., CEMATSL. D., SLLCKI, N., KAWAMURA, J., I-.Aw~L\-.AKI, T . , ITOH, N. & ORARA, K. 1985. Role of endothelium in responses of pial vessels to change '' in blood pressure and to carbon dioxide in cats. 3 Crrrb blood flom .Uetcib 7, suppl. I , S 275. I IARDER, D.R. 1987. Pressure-induced myogenic activation of cat cerebral arteries is dependent on intact endothelium. Ci,r Rrs 60, 102-107. I k R T % , 11.hl., IIEM.\lISGSEN, R., BOI.WIG, T.G. 1977. Rapid and repetitive measurements of blood flow and oxygen consumption in the rat brain using intracarotid xenon injection. .4ctu Ph,ysiol Srund 101, 501--503. K-ATLSIC, Z.S., SIIEPHERD, J.T. 81 V A w o w r E , P.M. 1985. Endothelium-dependent contraction to stretch in canine basilar arteries. .4in f Physiol. 252, €I 671-11 073. LASSES,N..-\. 196-1. Autoregulation o f cerebral blood no\\-. C i ~ Rf ~ 11/15 s (SUPPI. l), 201-204. P%LUER, R..\I.J., FERRIGE, A.G. & MONCADA, S. 1987. Nitric oxide release accounts for the biological activit! of endothelium-derived relaxing factor. .\.dtiw 327, -i2Cj26. P.ALLSOS, O.B., STRAKDGAARD, S. & EDVINSSON, L. 1990. Cerebral autoregulation. Crrebrowsr Brain .bfrtub Re? 2 , 161-192. SCt-l.\llDl, J.F., ~ ~ A L D E M A R G., , \'ORSTRCP, S., ASDERSEN,A.R., GERRIS, F. & PALLSON, O.B. 1990. Computerized analysis of cerebral blood flow autoregulation in humans : Validation of a method for pharmacologic studies. 3 Curdiorusc Phicrrnucol 6, 983-988. \V.ALG, (A, PALLSON, 0 . B . & LASSES,N.A. 1991. Is EDRF (NO) involved in hypercapnic cerebral \ asodilation ? In : SecxnJ Internutionul Meeting : Biolrtgy of'.\-ttrir Oxide, London, OCI 1-3, 1991. GOTOtl,
Is autoregulation of cerebral blood flow in rats influenced by nitro-L-arginine, a blocker of the synthesis of nitric oxide? Q WANG, "0.B. P A U L S O N and N. A. L A S S E N Department of Clinical Physiology and Nuclear Medicine, Bispebjerg Hospital, "Department of Neurology, Rigshospital, Denmark Autoregulation of blood flow is the term used to describe the constancy of organ blood flow observed when the arterial blood pressure is varied. It implies constriction of the resistance vessels, the arterioles, when the pressure increases and their dilatation when the pressure decreases. Autoregulation is seen in all organs of the body, but it has been studied in the cerebral circulation in particular (Lassen 1964, Paulson et al. 1990, Schmidt et al. 1990). Many theories have been proposed to explain the response. Most authors have favoured the myogenic theory. It holds that the smooth-muscle cells of the arterial wall are alone in eliciting the response. These cells are assumed to 'sense' the increased stretch caused by elevation of the blood pressure and to respond to this stretch by a constriction and vice versa. Recently, however, evidence has been published pointing to a key role of the arterial endothelium in autoregulation of CBF and probably also of blood flow through other organs. Studies on isolated segments of arteries from the brain of cats and dogs have shown that the normal increase in vascular tone to stretch is abolished if the endothelium is removed (Harder 1987, Katusic et al. 1987). In studies on the diameter of the pial arterioles of cats it was demonstrated that damage to the endothelium caused by ultraviolet light abolished the normal vasodilator response to a reduced blood pressure, while the same vessels have a preserved vasodilatation to hypercapnia (Gotoh et al. 1987). A number of vasoactive substances are now known to be produced by the endothelium. They comprise both potent vasoconstrictors and potent vasodilators. In particular the very rapidly cleared endotheliumderived relaxing factor (nitric oxide) (Palmer et al. 1987) might seem to be a suitable candidate, as it would explain the short time constant of the autoregulatory response. It is of interest, therefore, to Received 6 March 1992, accepted 30 March 1992. Key words : autoregulation, cerebral blood flow (CBF), nitric oxide (NO), nitro-1-arginine (NOLAG). Correspondence : Niels A. Lassen, Department of Clinical Physiology and Nuclear Medicine, Bispebjerg Hospital, DK-2400, Copenhagen NV, Denmark.
study cerebral autoregulation when nitric oxide (NO) synthesis is blocked. Nitro-L-arginine (NOLAG), a potent inhibitor of N O synthesis, was used in the present study. Its effect on CBF autoregulation was studied in anaesthetized (0.6% halothane/70% N,O), artificially ventilated Wistar rats. Cerebral blood flow (CBF) was measured by the intra-carotid '33Xe injection method as adapted for small rodents (Hertz et al. 1977).
50
I00 MABP (mmHg)
I50
. . . . . . . . . . . . . . . . . . . . . 50 100 I50 MABP (mmHg)
Fig. 1. Cerebral blood flow autoregulation in (a) control (saline) and (b) NOLAG-treated animals. CBF is given as a percentage of baseline values (in NOLAG group, the first measured CBF after NOLAG served as baseline). The autoregulation curve was drawn and low limits were calculated by a computerized procedure (see text).
297
298
Q. Mnng, 0. B . Paulson und R;. A. Lassen
The loner limit of CBF autoregulation \\as studied in 17 rats. Following measurement of baseline CBF, nine rats \\ere given 7.5 mg kg ' SOL.-\G. 0.1 ml min for h min intracarotidly. The remaining eight served as control, infusing only the vehicle (saline). In the control animals \I.IRP was raised gradual]!- to a level 20- 30 n m H g abo\-e baseline \l.IRP b! an infusion of noradrenaline (?---to x IO-" nig min I ) in order to study the expected plateau of CBF autoregulation. The blood pressure n a s then allowed to drop slowly by controlled bleeding t o the lovest measurdhk h e ! . CBF u a s measured a t approsimately 10 mmHg intervals of JLIBP during the whole stud!. In NOI.r\G-treated animals, the increase in blood pressure a a s noted. rendering it unnecessary to use noradrenaline. The autoregulation curl-e \\as calculated from the pooled CBF and .\l.iBP data in each group of animals, using a computerized procedure described else\\ here (Schmidt et ti(. 1990). Throughout the M hole experiment, bod! temperature, arterial PaO,, PaCO?, p€I were continuously monitored and maintained in the physiological range. There I\ ere no signiticant differences bet\\een saline and SOT. I G groups in this respect. The results showed that in all rats autoregulation of CBF \\-as preser\-ed (Fig. 1). NOI..\G (7.5 mg kg ') elicited an increase in \I.IBP h! .3h",, and a decrease in CBF b! 23",, as u e had seen in a previous stud!(\\'ang t t u/. 1991). .It this lon-ered CBF l e d the studied autoregulation of CBF was intact, as practicall! the same lover limit was observed in both groups, i 6 & 3.4 mmfIg for saline- and 78 6.1 mndlg for SOI,.-\G-treated animals (mean iSE, P > 0.05). In a pre\-ious stud!- we have shown, that the same dose i d N 0 1 , I G of 7..i mg kg \\-ill cause a marked suppression of the carbon dioside responsiveness of CHF (\\rang t! a/. 1991). The effects on SLIBP and CBF seen in this stud!- also constitute el-idence of the biological dctit-ity of the dose gi\en, effects that pe for at least 2 h. On this basis we conclude that 5: nthase actii-it! \\as indeed eKecti\-el!- inhibited in our studies. The tindings of a preseried CBF autoregulation consequentl! mean that this regulation is probahl! not dependent on endothelium-deriwd nitric oxide. Ltl'hen 30 mg kg ' NOLriG \\-as infused in our earlier stud! (M'ang el L i / . 1991), ~e observed an increase in blood pressure \$ hich exceeded the upper limit of(:HF autoregulation ( > 150 mmIIg) (Paulson r t nl. 1990). In some rats the blood pressure rose as high as 170 mmfIg, but CBF still remained constant (unpublished observation). Thus the autoregulator! plateau seems widened in the high-pressure range, as it does in the case of low CBF caused by hyperLentilation. .Is noted in the introduction, remoial of the
endothelium abolishes the normal constrictor response of isolated cerehral \-essels to stretch (Harder 1987, Katusic el izl. I%;), and destruction of the endothelium in a pial vessel abolishes the normal dilation response resulting from a drop in blood pressure (Gotoh et ctl. 1987). Our demonstration of preserved CBF autoregulation despite NO synthase blockage suggests, therefore, that the results mentioned ma! he due to elimination o f other endothelium-deriwd factors, rather than EDRF (NO). The stud?- was supported bl; the Alfred Benzon Foundation, the Lundbeck Foundation and the Danish Health Xledical Council, Denmark. The escellent technical assistance of Kim Pape Gadegdard and Pia Tejmer is gratefully acknowledged.
REFERENCES F., F L K W C I I I , y.,h I A N O , T., TANAK.4, K., CEMATSL. D., SLLCKI, N., KAWAMURA, J., I-.Aw~L\-.AKI, T . , ITOH, N. & ORARA, K. 1985. Role of endothelium in responses of pial vessels to change '' in blood pressure and to carbon dioxide in cats. 3 Crrrb blood flom .Uetcib 7, suppl. I , S 275. I IARDER, D.R. 1987. Pressure-induced myogenic activation of cat cerebral arteries is dependent on intact endothelium. Ci,r Rrs 60, 102-107. I k R T % , 11.hl., IIEM.\lISGSEN, R., BOI.WIG, T.G. 1977. Rapid and repetitive measurements of blood flow and oxygen consumption in the rat brain using intracarotid xenon injection. .4ctu Ph,ysiol Srund 101, 501--503. K-ATLSIC, Z.S., SIIEPHERD, J.T. 81 V A w o w r E , P.M. 1985. Endothelium-dependent contraction to stretch in canine basilar arteries. .4in f Physiol. 252, €I 671-11 073. LASSES,N..-\. 196-1. Autoregulation o f cerebral blood no\\-. C i ~ Rf ~ 11/15 s (SUPPI. l), 201-204. P%LUER, R..\I.J., FERRIGE, A.G. & MONCADA, S. 1987. Nitric oxide release accounts for the biological activit! of endothelium-derived relaxing factor. .\.dtiw 327, -i2Cj26. P.ALLSOS, O.B., STRAKDGAARD, S. & EDVINSSON, L. 1990. Cerebral autoregulation. Crrebrowsr Brain .bfrtub Re? 2 , 161-192. SCt-l.\llDl, J.F., ~ ~ A L D E M A R G., , \'ORSTRCP, S., ASDERSEN,A.R., GERRIS, F. & PALLSON, O.B. 1990. Computerized analysis of cerebral blood flow autoregulation in humans : Validation of a method for pharmacologic studies. 3 Curdiorusc Phicrrnucol 6, 983-988. \V.ALG, (A, PALLSON, 0 . B . & LASSES,N.A. 1991. Is EDRF (NO) involved in hypercapnic cerebral \ asodilation ? In : SecxnJ Internutionul Meeting : Biolrtgy of'.\-ttrir Oxide, London, OCI 1-3, 1991. GOTOtl,
