Blomer, 1955). Very little is known about the control of deep cerebral blood

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1 1 J. Physiol. (I956) I33, IO-3 FACTORS INVOLVED IN THE CONTROL OF CEREBRAL BLOOD FLOW BY A. CARLYLE AND J. GRAYSON From the Department of Physiology, University of Bristol and the Department of Physiology, University College, Ibadan (Received 29 November 1955) Most previous knowledge of the cerebral circulation was based on the measurement of blood flow in the carotid arteries (Rein, 1929) or pial blood vessels (Forbes, 1928), or on the measurement of total brain blood flow by clearance methods (Kety & Schmidt, 1945) or radioactive methods (Nylin & Blomer, 1955). Very little is known about the control of deep cerebral blood vessels. In the present work blood flow changes were recorded in the substance of the frontal lobes by means of 'internal calorimetry'. This is a development of the heated thermocouple method (Gibbs, 1933), which was originally applied qualitatively to brain blood flow studies by Schmidt & Pierson (1934). In its present form, the technique gives a record of thermal conductivity, which in other organs has been shown to be a linear function of blood flow (Grayson, 1952). The first object of the present work, therefore, was to demonstrate the validity of the method for the brain. The second object was to use it first to determine the importance of neural factors and adrenaline in the control of deep cerebral blood flow, and secondly to investigate the adaptations which take place in the intracerebral circulation during changing blood pressure. METHODS A development of the technique of 'internal calorimetry' (Grayson, 1952) was used for the measurement of thermal conductivity in the brain. The recording instrument is a copper-constantan thermocouple with an attached filament of constantan through which a heating current can be passed. It has been shown (Grayson, 1952) that, when a constant current is passed through the heater, heat exchanges in such an instrument are governed by the simple relation J2=Fk (where I= heating current, =temperature increment, k = thermal conductivity of medium, F = instrumental factor). F is constant for any given recorder and its determination has already been described.

2 CEREBRAL BLOOD FLOW From the formula, 'k' can be calculated if we know '1' and ''. In the practice of 'internal calorimetry', as previously described, the current required to produce a temperature rise of 1C was determined using a manually operated apparatus and visual means of estimating temperature and current. An element of personal skill was involved and there was a possibility of personal error, particularly when the base-line temperature was subject to change. In their muscle blood-flow recorder Hensel, Ruef & Golenhofen (1954) solved the difficulties by using a large needle incorporating heater, thermocouple and cold junction. This was not applicable to the present work, partly because the instrument was too large, partly because its working depends on the assumption that tissue temperature is the same both at the recorder and at the cold junction. Since Gerard & Serota (1936) showed that sharply localized temperature changes could occur in the brain, this assumption could not be regarded as valid for our present purposes. In the present work, therefore, a different development of the technique was used to eliminate personal factors. Temperature was recorded continuously on photographic paper by means of a moving coil galvanometer, slit lamp and recording camera. Fixed values of la were supplied to the heater by an automatically operated device and the corresponding temperature elevation measured on the photographic record. As the equation shows, k was an inverse function of the temperature increment,, and could be calculated. A complication in the development of this method of recording was the fact that on applying current to the filament 7 min was needed for the attainment of temperature equilibrium (Grayson, 1952); again on switching off the current 7 min was required for full cooling. Readings could not, therefore, be taken at less than 14 min intervals. To cut down the time necessary, a system of overheating and overcooling was adopted. This was made possible by beginning the readings from an already elevated temperature base-line (lower equilibrium temperature) produced by a current of -447 A (12=-2). The temperature corresponding to a further J2 increment of -115 was termed 'upper equilibrium temperature'. In order to obtain stable upper and lower equilibria, current was supplied to the heater as follows: (1) -447 A (12=-2) for 24 sec lower equilibrium; (2) -775 A (12 =-6) for 17 sec; (3) -561 A (12 =-315) for 24 sec upper equilibrium; (4) zero current for 17 sec. The cycle thus incorporated short bursts of overheating and overcooling with 24 sec at each equilibrium level. The difference between the upper equilibrium and lower equilibrium temperature was thus the temperature rise produced by an 12 increment of -115 and k could be readily calculated from the equation. The current and time values were arrived at empirically. It was important that equilibration should always be complete over a wide range of thermal conductivity. Accordingly, tests were made on inert media of thermal conductivity ranging from 11-2 x 1-4 to x 1-4 c.g.s. units. Results obtained by this method and by the manual method were identical throughout the range. Apparatus. The apparatus used to deliver heating current according to the above cycle is illustrated diagrammatically in Fig. 1. The source of current was a 1 A-hr 6 V accumulator. M1, M2 and M3 were microswitches operated through cams driven by a constant speed motor. At one position of the cams all three switches were open so that no current passed. Each heater lead contained a fixed resistance, R1, R2, R. to bring the current in each lead within the required range. Fine control was effected by the slide wire resistances in the switching box (Fig. 1 B). An additional -1 Ql decade resistance was included in the lower equilibrium circuit, since with low currents the -1 Q1 slide wire resistance would not give sufficient flexibility. Heater resistance varied slightly from recorder to recorder. Accordingly, a Cambridge L.Y. unipivot galvanometer was connected to the main heater circuit through a vacuojunction. This enabled 12 values to be read directly, and adjustments to be made at the beginning of each experiment by means of the variable resistances. The switching box shown in Fig. 1 B housed the adjustable resistances and the microammeter, it also incorporated a circuit whereby a standard manual box (Grayson, 1952) could be brought into use to check the 12 value at any one time. Wiring details will be apparent from the figure. ii1

3 12 A. CARLYLE AND J. GRAYSON To facilitate adjustment of the recording light beam on the camera scale a simple potentiometer circuit as shown in Fig. 1C was used. This was sufficient to compensate for a 15C difference in temperature between cold junction and recorder. The recorders themselves were of the standard pattern described elsewhere, with a slight modification that 36 s.w.g. copper and constantan wire was used for the thermocouple, the constantan wire being prolonged 4 mm with 42 s.w.g. copper wires (heater leads) soldered at the tip and 1 mm from the thermojunction-making the heater filament 3 mm long. A R - U ~~~T4 -f m 2 R2, T3fie c a r aaa B T, T,cneinfomthe cam operated supply.(b).sw vitg. A :T2o-r 3 3 T4 To recorders A ~~~15.1.*nc * 1.t 1 15VH -a~~~~~~~~~~~~~~~- Houple C2 Ci R G I'~~~~~~~~~~~~~~~~~ Fig. 1. Internal calorimetry. Wiring of ancilary apparatus for photographic recording. (A) B, 1eA-hr 6rV accumulator. Me, M2, M.c, microswitches operated by cam from constant speed motor. R,mal, 14, fixed calibrating resistances to give approximate output currents. TlȚ2m T3, T4, connexions from the cam operated siupply. (B). Switching box. V.J., vacuojunction with output connected to Cambridge L.Y. microammeter, A, calibrated by variable resistance to reade odirectly. Y, Yaxley switch shown connecting switches to heater leads H2 and completing the thermoelectric circuit from recorder 2 through the junction box J.B. to the cold junction. In the alternative position of the Yaxley switch heater leads HI from the manual box are connected to the recorder heater leads H2; the recording thermocouple 2is connected through the junction box to leads Cl which connect to the manual box with its independent cold junction.g, connexions to the recording galvanometer. R, external resistance in galvanometer circuit. C.J., cold junction leads. (C). Potentiometer to introduce small opposing currents into thermoelectric circuit for purposes of scale adjustment. Thermal conductivity and brain blood flow. The application of the present technique to bloodflow measurement is exactly the same as previously described for 'internal calorimetry' (Grayson, 1952). It has been shown that in liver (Grayson, 1952) and spleen (Hensel et al. 1954) over the biological range, thermal conductivity is an approximately linear function of blood flow. It might be reasonable to suppose that the same would also apply in the brain. However, perfusion experiments were carried out to investigate this. A blood-flow recorder was implanted in the frontal lobe of a lightly anaesthetized cat. The vertebral arteries and veins were ligated, the

4 CEREBRAL BLOOD FLOW 13 femoral arteries cannulated and connected to a reservoir of blood, itself maintained under controlled pressure. All jugular veins were cannulated headwards (the number of jugular veins varied in different animals but usually there were two main internal jugular and two main external jugular veins). Arterial branches supplying the muscles of the neck and face were ligated as far as possible. The blood flowing down the jugular veins was collected and the rate of flow measured by means of measuring cylinder and stop-watch. The blood was then returned by means of a pump, via the reservoir, to the systemic circulation. It was probably reasonable to assume that the venous outflow measured was very largely cerebral outflow x 7 -c E Rate of blood flow (ml./min) Fig. 2. Graph of conductivity increment (in c.g.s. units), recorded in the frontal lobe of the cat, against jugular venous outflow, showing a straight-line relationship over the physiological range. Arterial pressure was lowered in stages to zero and thermal conductivity of the frontal lobes compared with venous outflow. It will be seen from Fig. 2 that the correlation was approximately linear. Similar results were obtained in all such experiments. Implantation of blood-flow recorder8. In the present experiments cats, rabbits, sheep and kid goats, lightly anaesthetized with pentobarbitone sodium (Nembutal, Abbott Laboratories) were used. The skull was exposed by a longitudinal incision and a trephine hole placed, just lateral to the mid-line and i in. anterior to the level of intersection of lines from the mesial canthus of the eye to the base of the contralateral ear, in the case of the cat and rabbit, and at the level of the intersection of lines from the mesial canthus of the eye to the base of the contralateral horn in the sheep and goat. Under direct vision the recorder was inserted through the dura to a depth which varied with the species. In the rabbit this was about 2 cm, the direction of insertion being such as to bring the recorder tip near the centre of one or other frontal lobe. The trephine hole was packed loosely with cotton-wool and the recorder further secured by a suture through a loop in the wire and by the closure of the skin incision around it. Blood-pressure recording. In the rabbit blood pressure was recorded photographically from the femoral artery (the rabbits were fully heparinized). In sheep and goats, mercury manometers and kymograph recording were used and clotting was avoided by a continuous slow mechanical infusion of saline through the recording cannula.

5 14 A. C(ARLYLE AND J. GRAYSON Blood-pres8ure compensation. In order to impose blood-pressure changes in fully heparinized rabbits and cats, the other femoral artery was cannulated and connected to a reservoir of donor blood. The reservoir was connected to a large bottle in which the pressure could be regulated by means of a foot pump and manometer. On lowering the pressure in the pressure bottle, blood flowed from the animal into the reservoir until equilibrium was attained. Arterial blood pressure could thus be lowered from resting level to zero. The compensator was also used to prevent blood-pressure change during an adrenaline infusion. With a cannula in the femoral artery, however, the exchange between the reservoir and the animal was not sufficiently quick, and in a few experiments a common carotid artery was used. RESULTS Thermal conductivity was recorded from the frontal lobes of sheep, goats, rabbits and cats. In twenty-four experiments 12 values (see Methods) were measured with apparatus operated manually, in nineteen experiments the apparatus was automatically operated, photographic recording being used as described. There was no significant difference between the results obtained by these two methods. Photographic recording was, of course, easier in operation; it was possible with this method to analyse small changes with greater accuracy and to allow for changes in tissue temperature. The technique was particularly valuable with small blood flows. Resting variations in anaesthetized animals A wide scatter of blood-flow readings has been obtained throughout the present work. Conductivity increments recorded in sheep, lightly anaesthetized with pentobarbitone sodium, ranged from 4-6 to 15-1 x 14, with mean values of 7-6 x 14. In rabbits the range was 2- to 15-2 x 14, with a mean value of 6-5 x 1-4, in goats, 5-9 to 11-4 x 14, in cats 6- to 9- x 14, with a mean value of 7-8 x 1-4. In one animal the same recorder was re-inserted four times and conductivity increments of 1, 3, 3 and 12 x 14 were obtained from these slightly different sites of implantation. Variations between different sites of implant were observed in many different experiments. Yet in an animal at rest readings of 8k from any individual implant were constant and showed no fluctuation. Effect of nerve block on resting blood flow in the brain Direct denervation of the cerebral blood vessels was not attempted, but ganglion blocking agents, hexamethonium bromide and tetraethylammonium chloride were used for the same purpose. Fig. 3 illustrates a typical finding. In this experiment on one animal photographic recording was used to calculate thermal conductivity in the rabbit brain, the animal being lightly anaesthetized with pentobarbitone sodium. Fig. 3A shows a section of the record when blood pressure was 1 mm Hg, 6 was.415 C, whence the thermal conductivity was 25-2 x 14. Thermal conductivity recorded by the same

6 CEREBRAL BLOOD FLOW 15 instrument after death, calculated from the section of the record shown in Fig. 3D, was 1 x 14, whence by subtraction the excess thermal conductivity, 8k, due to circulating blood, was 15X2 x 1-4. The record shown in Fig. 3B was taken after a total dose of 3 mg tetraethylammonium chloride had been administered in three doses of 1 mg at 1 min intervals (the last dose had no effect on blood pressure). Blood pressure after 'ganglion blocking' in this way was 1 mm Hg; 8k was calculated to be 12-8 x 1-4. A further dose of 2 mg atropine was given and the effect is shown in Fig. 3 C. Blood A Control B + TEAC B C C + Atropine D- Dead 2-1 U ) 1-5 to -C C f 1. I 4 L. co E l- B.P 1mm Hg =-415 C k=25-2 x1-4 B.P. 1mm Hg =-46 C k=22-8 x1-4 B.P 94 mm Hg 9 =45 C k=23-1x1-4 B.P. mm Hg C k=1x I - I ) Time (min) Fig. 3. Internal calorimetry: photographic record of the temperature of a heated thermocouple in the frontal lobe of a rabbit. The heating current to the thermocouple was automatically varied in a cyclical manner as described in the text. (A) Control: animal anaesthetized with Nembutal. (B) After ganglionic blockade with tetraethylammonium chloride (TEAC). (C) After administration of atropine. (D) Animal dead. Values of k calculated from photographic trace using the equation k= 115/OF (=temperature difference at the two equilibrium levels; F for this instrument =11). pressure was 94 mm Hg and 8k was now 13-1 x 1-4. That this was a sensitive preparation was shown by lowering the blood pressure transitorily below 4 mm Hg; 8k fel to 6-6 x 1-4, but recovered on restoring the blood pressure to 9 mm Hg. In this experiment then, administration of tetraethylammonium chloride had no significant effect on the resting level of cerebral thermal conductivity.

7 16 A. CARLYLE AND J. GRAYSON Similar results were obtained in eleven experiments. Atropine was given in twelve experiments. This substance, too, had no significant effect on the equilibrium level of cerebral thermal conductivity. Action of adrenaline on cerebral bloodflow Fig. 4 shows the result of a typical experiment on a rabbit in which an intravenous infusion of adrenaline hydrochloride was administered at the rate of 5,ug/min. Blood pressure rose rapidly from 9 to 148 mm Hg and bo I 17 E El, co - V I x C> U- -o Adrenaline (5,tg/min) Fig. 4. Effect of intravenous infusion of adrenaline on cerebral blood flow in the rabbit, showing record from heated thermocouple in frontal lobe, femoral arterial pressure and, below, conductivity increment (8k) calculated from record. remained at this level throughout the period of the infusion. Sk, calculated from the photographic trace, rose at the beginning of the infusion from 5 x 1-4 to 9.1 x 1-4. The maximal elevation of flow indicated by the 8k value was recorded 11 min after the beginning of the infusion. Thereafter blood flow declined slowly during the period of the infusion. During the same period there was a simultaneous fall in brain temperature. On stopping the infusion blood flow fell just below the resting level, returning again within 3-5 min. Thirty-six adrenaline infusions were given to twelve rabbits in doses varying from -5 to 1lg/min. Results were qualitatively similar in all cases.

8 CEREBRAL BLOOD FLOW 17 Similar results were obtained in sheep, goats and cats. Thus, provided the dose of adrenaline was sufficient to raise the blood pressure, cerebral 5k values rose to a maximum level within 3 min, and thereafter in most experiments declined towards resting levels, even though the infusion was continued and the blood-pressure elevation maintained. In long continued infusions (up to 2 min at 5pg/min in the rabbit) 3k never dropped below resting levels during the period of infusion, but on stopping 3k was occasionally observed to fall transiently below resting levels. L 15 AA I E E15 ~~~~~~ 9~~~~~~~~~~~~~~~ x L A L --- Jco Time (min) 2<g x3 I ~~~~~~~~E5 Time (mm) C 2 Adrenline 5 /igmin) Adrenaline (5,ug/min) Fig. 3. Effect of intravenous infusion of adrenaline, showing photographic record and calculated 6k. (A) Rabbit anaesthetized with Nembutal, no blood-pressure compensation. (B) Systemic arterial pressure compensated. Adrenaline infusion following blood-pressure compensation Fig. 5 B shows the results of an experiment in a rabbit in which blood pressure was prevented from changing by connecting the compensating reservoir to a carotid artery (compensation through the femoral artery was ineffective). In this case the cannula was inserted headwards, and blood pressure recorded from a femoral artery. On occasions when femoral arterial pressure showed very little change during intravenous adrenaline infusion it may, perhaps, be assumed that there was even less change in the effective perfusion pressure of the brain. Adrenaline in a dose of 5,ug/min infused into the femoral vein, before compensation (Fig. 5 A), produced a 73% rise in blood pressure and a 1 % increase in 3k; after compensation the same dose produced a decrease of 7 % in 8k. In a second experiment the compensation was less effective, nevertheless, the blood-pressure response was greatly reduced, and a dose of adrenaline 2 PHYSIO. CXXXIII

9 18 A. CARL YLE AND J. GRAYSON which had previously caused a 6% increase in blood flow now produced only a 1% increase. The effect of intracarotid infusions of adrenaline. Infusion of adrenaline direct into the carotid artery was carried out in eight experiments on different animals (Table 1). Doses were similar to those previously used intravenously but the results were significantly different. Whereas in nearly every intravenous adrenaline infusion Sk rose by 4-8 %, in 3 min (see Tables 2 and 3), in only one intracarotid infusion was there a slight rise (5-5 %) in ak. In the other experiments 8k fell by significant amounts. TABLE 1. Effects of intracarotid infusions of adrenaline 8k After After 5 mim Infusion 3 min 7 min after end rate Animal Resting infusion infusion of infusion (yg/min) Goat 7* Goat 8*2 6X 8X 8-1f Sheep 7* *2 7-2 Sheep Sheep Rabbit Rabbit Rabbit f 5 TABLE 2. The effects of intravenous adrenaline infusion before and after administration of ganglion blocking agents. In each case blood-pressure change after blocking equalled or was slightly greater than that before. HMB =hexamethonium bromide; TEA = tetraethylammonium chloride Percentage flow increase after 3 min infusion Animal Agent Before block After block Goat HMB Goat TEA Goat TEA +8, +5 Sheep TEA Rabbit TEA Rabbit TEA + 6 Rabbit TEA TABLE 3. The effects of intravenous adrenaline infusion before and after administration of atropine. In each case blood-pressure change after atropine equalled or was greater than that before Percentage flow increase after 3 min infusion Animal Before atropine After atropine Goat Goat Goat Rabbit Rabbit

10 CEREBRAL BLOOD FLOW 19 During the first 3 min of such an infusion there was usually little change in pressure in the femoral artery; thereafter it usually rose. In some experiments it was similar in extent to that produced by the same dose of adrenaline administered intravenously. In four of the experiments where the blood pressure rose, an increase in cerebral 8k values also occurred. Such an increase was never seen until the blood pressure had risen. The effect of ganglionic blocking agents on cerebral responses to adrenaline. In eighteen experiments on sheep, goats and rabbits, intravenous infusions of adrenaline sufficient to cause blood-pressure increments of 5-1% were accompanied by characteristic short-lived elevations of 8k. Administration of tetraethylammonium chloride profoundly modified the effect. The seven results given in Table 2 were selected because in each case, after an effective blocking dose, the blood-pressure response to a 5 min infusion was, after 3 min, the same or greater than the response before blocking. In spite of the occasionally enhanced blood-pressure response, the 8k response to adrenaline infusion was in all cases diminished and in some cases reversed. The effect of atropine on blood-flow responses to adrenaline. The results of five experiments are given in Table 3. These cases were selected from a wider series because, after atropine, the blood-pressure responses to a fixed dose of adrenaline were the same or slightly greater than before. In four instances the brain blood-flow responses were reversed, in the other the effect of a given dose of adrenaline was greatly decreased. Carotid arterial contribution to cerebral blood flow Clamping one carotid artery alone never produced any lasting effect on cerebral 8k values in sheep, goats or rabbits. In most experiments 8k increased transiently, with the rise in blood pressure which followed carotid clamping. Clamping both carotids produced variable effects. Fig. 6 shows two typical results. The experiment shown in Fig. 6 A was performed 2 hr after insertion of a cannula, directed headwards, into the left carotid artery. Pressures in the cranial stump of the carotid artery could thus be recorded. On clamping the remaining carotid artery, systemic arterial pressure rose, pressure in the carotid stump fell, then slowly returned to the resting level. Simultaneously there was a rise in cerebral blood flow which was not sustained. Similar results were obtained in all such experiments except that the blood flow changes were frequently less marked. Fig. 6B shows the effect of sudden clamping of both carotid arteries in the sheep. Headward pressures were not recorded. Blood flow fell rapidly then slowly recovered until, after 15 min, it was practically at resting level. Similar results were obtained in all such experiments. In both types of experiment it is interesting to observe that after 15 min clamping, brain blood flow had always returned to the resting level. 2-2

11 2 A. CARLYLE AND J. GRAYSON 4) A o _ boe x -- co ICD 5 54o _ ~~ ~ ~~~~~~~~~dk r_ 3 -Head pressure -165L. s 2 - left carotid B15 O. A B k13 12,_ -- > j Femoral pressure Femoral pressure A 5 1tClamp on right carotid IClamp on both carotids 6 4 so l; Time (min) Fig. 6. Effect of carotid occlusion upon cerebral blood flow of the sheep. (A) Conductivity increment (8k) in frontal lobe, blood pressure in femoral artery and in cranial stump of divided left carotid artery during occlusion of intact right carotid artery. (B) Conductivity increment in frontal lobe and femoral arterial pressure during simultaneous occlusion of both intact carotid arteries. The effect of changing blood pressure on cerebral blood flow During the performance of a large number of experiments involving the administration of adrenaline, etc., it was observed that although systemic blood pressure itself fluctuated considerably during the course of an experiment, there was little change in the resting level of cerebral blood flow in most animals. Accordingly, experiments were carried out to investigate the relation between systemic arterial pressure and blood flow in the brain. Cats and rabbits, lightly anaesthetized with Nembutal, were used; blood pressure was recorded in a femoral artery. The other femoral artery was connected, through as wide a cannula as could be inserted, to the blood-pressure compensator. 8k was calculated for the frontal lobe. In a few experiments an external jugular vein was also cannulated for the recording of venous pressure. Fig. 7 shows the results of a typical experiment on a cat, in which systemic arterial blood pressure was lowered in stages from 93 mm Hg to zero. Cerebral 8k values showed no significant changes in level until the blood pressure reached 4 mm Hg; thereafter they declined rapidly. In most experiments, each rapid drop in blood pressure produced a transient fall in 8k; when the blood pressure was maintained at its new level, 8k always returned to its previous level within 5 min. Fourteen experiments of this kind were performed on the rabbit. Fig. 8 A shows a similar experiment. In this experiment there was a slow decline in flow with blood pressures below 6 mm Hg. Blood flow at 3 mm Hg >U. 4)o I

12 CEREBRAL BLOOD FLOW x 4)i L_ E di 4 :L 4., 3 -._ 3ve 2 1 Fig Time (min) Graph of conductivity increment (8k) and systemic blood pressure in the anaesthetized cat against time, the blood pressure being lowered in stages to zero Arterial blood pressure (mm Hg) Fig. 8. Effect of ganglionic blockade upon blood-flow response to lowered systemic arterial blood pressure. Graph of conductivity increment in the frontal lobe against femoral blood pressure. (A) Rabbit anaesthetized with Nembutal. (B) After effective ganglionic blockade with tetraethylammonium chloride.

13 22 A. CARLYLE AND J. GRAYSON was 25 % less than that at 7 mm Hg. Maximal change occurred between arterial pressures of 3 and 2 mm Hg. When the arterial pressure was 15 mm Hg cerebral blood flow was not measurable. The effect ofganglion blocking agents on the blood-pressure responses. Systemic arterial blood pressure in five rabbits was lowered in stages after the administration of tetraethylammonium chloride. Fig. 8B shows a typical experiment of this kind. The form of the curve is similar to that seen in Fig. 8A. XZ AX It_ A Time (min) Fig. 9. Reversibility of cerebral blood-flow response in the rabbit to lowered systemic arterial blood pressure; graph of conductivity increment in the frontal lobe and femoral arterial blood pressure against time. (A) Arterial blood pressure lowered in stages to 1 mm Hg and then raised. (B) Arterial blood pressure reduced rapidly (at arrow) to 1 mm Hg and then raised slowly. 8k began to decline when the arterial blood pressure fell below 75 mm Hg, the maximal fall occurring when the pressure fell below 3 mm Hg. 8k was not measurable when the arterial pressure was 15 mm Hg. In all similar experiments the equilibrium levels were not affected by ganglion blocking, but the transient falls in blood flow, which, in the unblocked animal, occurred with each blood-pressure decrement, were less apparent. In three of the five experiments, indeed, they were completely absent. In the other two exmperiments they were small. Reversibility of the blood-pressure response. Fig. 9A shows the result of an experiment on a rabbit in which the blood pressure was lowered in stages to

14 CEREBRAL BLOOD FLOW 23 1 mm Hg. It will be seen that with the blood pressure at 3 mm Hg, 8k was about 25 % lower than when the blood pressure was 65 mm Hg. When the blood pressure was lowered to 18 mm Hg, blood flow declined to zero. In this experiment blood pressure was successfully maintained at levels below 18 mm Hg for about 8 min. The blood which had passed into the compensator was then returned to the animal and blood pressure was restored to 55 mm Hg. Cerebral blood flow recovered to its initial resting level. Similar results were obtained on two other occasions. In most experiments, however, keeping the blood pressure below 2 mm Hg for more than 2 or 3 min caused the death of the animal. A potent cause of death was respiratory failure, and all the successful experiments involved the use of artificial respiration (by means of a standard pump and tracheal cannula). Even so, many animals died after a very short exposure to these extremely low blood pressures. Fig. 9B shows the results of an experiment in which the blood pressure was lowered rapidly from 75 to 15 mm Hg. Blood flow also fell rapidly to levels just above zero. In this experiment it was possible to restore the blood pressure slowly. It will be seen that, during this recovery phase, when the blood pressure was 25 mm Hg, blood flow had recovered to 5% of its initial value; with the blood pressure at 35 mm Hg, blood flow recovery was almost complete. Changes in cerebral vascular resistance during imposed low blood pressure. In three experiments jugular venous pressure was measured by inserting a widebore hypodermic needle into an external jugular vein and connecting it to a saline manometer. Fig. 1 shows the result of a typical experiment in the rabbit in which systemic arterial blood pressure was lowered in stages from 9 mm Hg to zero. It will be seen that venous pressure fell concomitantly. When the arterial pressure was 16 mm Hg, Sk was zero. Venous pressure at this stage was so low as to be almost unmeasurable. It is clear, however, that there was a positive perfusion pressure. This suggested that at low arterial blood pressures there was an active increase in cerebral vascular resistance. Accordingly, in three experiments cerebral resistance was calculated, on the basis of Ohm's law, in arbitrary units, as 'perfusion pressure'/'flow', i.e. systemic arterial pressure less external jugular venous pressure, divided by 8k. A typical result is shown in Fig. 11. It is clear that the terminal fall in cerebral blood flow, when blood pressure dropped below 2 mm Hg, was accompanied by a big increase in vascular resistance in the brain. Similar findings were obtained in each case. The effect of venous cannulation on the brain blood-flow responses to lowering of the blood pressure. In three experiments on rabbits, where it was desired to record venous pressure, an end cannula was inserted into one of the main external jugular veins. It was thought at the time that venous anastomosis

15 24 A. CARLYLE AND J. GRAYSON 4 I 3 E E 2 u 2, n> u Time (min) Fig. 1. Graph of conductivity increment in the frontal lobe of the rabbit, jugular venous pressure and femoral arterial pressure against time; the systemic arterial pressure being lowered in stages to zero. I 1-1 E E 7o x 4-6 'C> c D. 4, V._i, 3 no u Time (min) Fig. 11. Graph of conductivity increment in the frontal lobe of the rabbit, femoral arterial blood pressure and intracerebral vascular resistance against time. Vascular resistance plotted on arithmetical scale in arbitrary units as defined in text. When systemic arterial pressure reached 15 mm Hg, vascular resistance approached infinity, flow being zero.

16 CEREBRAL BLOOD FLOW 25 in the neck was so free that this procedure could scarcely interfere with the venous drainage, but would afford a more reliable and rapid means of measuring venous pressure than was possible with a hypodermic needle. The results of all three experiments were similar. Venous pressure changes were similar to those shown already in Fig. 1. Blood-flow responses, however, were profoundly modified. A typical result is shown in Fig. 12. Lowering the arterial pressure from 9 to 8 mm Hg produced an immediate drop in blood flow. Cerebral blood flow decreased with each fall in arterial blood pressure, nor was there any subsequent recovery. In these experiments the equilibrium levels of blood flow were linked with the level of arterial blood pressure. 6- E 3o. ul Blood pressure (mm Hg) Fig. 12. Effect of venous cannulation upon cerebral blood-flow response to lowered systemic arterial blood pressure. Graph of conductivity increment in the frontal lobe against femoral arterial pressure in a rabbit with an end cannula in one external jugular vein. DISCUSSION Distribution of cerebral blood vessels. A feature of the present investigation was the wide variation in conductivity increment (8k) in different parts of the frontal lobe, which could be easily detected simply by altering the position of the recorder. The significance of this variation is hard to determine, since with a recorder in a fixed position, even though the resting value of 8k was high, little further change occurred without specific stimulus. The method of implantation used so far has not permitted precise location of the recorder; it might have been in either the white or the grey matter, which may to some extent account for the wide variation in results. It is clear that no attempt can be made to evaluate the present figures in terms of total blood flow in the brain. It is, however, interesting to note that in any one cerebrum, areas exist in which the 8k value is of the order of 2x 1o-4. In the liver this would correspond to a blood flow of about

17 26 A. CARLYLE AND J. GRAYSON 15 ml./1 ml. of tissue/min. On the other hand, there are areas in which the Sk value is only 2 x 1-4, which in the liver would correspond to a blood flow value of only 15 ml./1 ml./min. There is a clear need for a fuller investigation of the distribution of blood vessels, preferably using animals with bigger brains than the rabbit, and using better methods to allow precise localization of the recorder. Despite this weakness in the technique employed in this investigation it has been possible to obtain consistent patterns of response. Control of cerebral blood flow. In recent years, opinion concerning the control of the cerebral circulation has undergone considerable change. The Monro-Kellie doctrine was that since the skull was rigid and its contents allegedly incompressible, vasodilator or vasoconstrictor mechanisms were in consequence impossible. The only effective controller of cerebral blood flow remaining was, therefore, the mean systemic arterial blood pressure (Kellie, 1824). To-day this view is no longer fully tenable, for many workers have demonstrated beyond doubt that vasomotor mechanisms exist in the brain. The mass of work, far too extensive for review here, is such that there can be little doubt that both sympathetic and para-sympathetic nerves exert their characteristic effects just as much on the brain blood vessels as on blood vessels elsewhere in the body. The present work supports this view in so far as it provides evidence that rapid increments in blood pressure produce vasodilatation through cholinergic mediators, acute drops in blood pressure produce vasoconstriction, probably through adrenergic mediators. Nevertheless, in the literature, there is considerable confusion as to the effects of sympathetic nerve section. Thus some workers have shown that cervical sympathectomy produces an increase in cerebral blood flow (Forbes & Wolff, 1928). Others, such as Ludwigs & Schneider (1954), using a heated thermocouple, showed that no change in cerebral blood flow followed section of the cervical sympathetic. In the present experiments the effect of cervical sympathectomy was not investigated. However, ganglion blockage was found to have no lasting effect on the level of blood flow in the brain. Larger doses of blocking agent might have had some effect. Possibly the techniques of measurement were inadequate to demonstrate small changes in flow, but it seems most reasonable to regard the present experiments as supporting the conclusions of Ludwigs & Schneider (1954). It may be that blood vessels in the deep substance of the cerebrum behave differently in this respect from the pial blood vessels. Certainly our results support the view that nervous factors do not in themselves determine the level of resting intracerebral blood flow. The action of adrenaline on brain blood flow. Forbes & Wolff (1928) showed without doubt that adrenaline, applied locally, caused constriction of the pial vessels. But the effect of systemically administered adrenaline on brain

18 CEREBRAL BLOOD FLOW 27 blood flow is more complex. In moderate doses, intravenous adrenaline was shown by Cushing (192) and Wiggers (1914) to diminish blood flow in the brain. In the spinal cord, too, Field, Grayson & Rogers (1951) obtained evidence of vasoconstriction in response to single intravenous injections of adrenaline. In bigger doses, however, the effect of adrenaline is more variable. Thus Wiggers (1914) and Schmidt (1934) showed in perfusion experiments that if the blood pressure rose appreciably, the cerebral blood flow also rose, owing partly to intracerebral vasodilatation. In the present experiments, too, the extent of the blood-pressure response was found to be important. In all species tested a continuous infusion of adrenaline in pressor doses produced an initial rise in blood flow, followed after 3 or 4 min by a slow return to resting levels. The initial rise was not seen in experiments where the blood pressure was prevented from rising, and the results of intracarotid infusion leave little doubt that the direct action of adrenaline on vessels of the frontal lobe is vasoconstrictor. However, the present experiments suggest that the blood-flow rise occurring at the beginning of an infusion is not a simple mechanical effect of the raised blood pressure, as has usually been assumed in the past, since the administration of ganglion blocking agents modified the response, occasionally reversing it and in all other cases diminishing it. These results suggest that there is a reflex component in the blood-flow response to adrenaline. This is supported by the finding that atropine had a blocking action on the adrenaline dilator response comparable with that of the ganglion blocking agents. The available evidence thus suggests strongly that although adrenaline locally applied to the brain blood vessels causes vasoconstriction, intravenously it produces a rise in cerebral blood flow dependent on the elevated blood pressure: this increases cerebral blood flow partly through the increased perfusion pressure and partly through cholinergically mediated intracerebral vasodilatation. Intrinsic regulation of cerebral blood flow. Many workers still regard arterial blood pressure as the main determinant of cerebral blood flow. Experiments in which carotid blood flow was measured during a fall in systemic blood pressure showed a decline proportionate to blood pressure (Ngai & Nelson, 1954). Our own experiments on animals with equally extensive cannulation of neck blood vessels produced similar results. Nevertheless, our experiments on animals where the operation was limited to a small trephine hole in the skull produced a completely different picture. There was no longer a correlation between blood pressure and blood flow; blood flow was substantially unaltered even by relatively large changes in systemic blood pressure. These findings, in our opinion, can only be interpreted as evidence of an additional regulatory mechanism in the brain, namely 'intrinsic regulation'.

19 28 A. CARLYLE AND J. GRAYSON The phenomenon of 'intrinsic regulation' was first described with reference to the kidney by Shipley & Study (1951). They showed that over a wide range of blood-pressure variation, kidney blood flow remained constant, falling significantly only when the systemic blood pressure fell below a critical value. This power of the renal vasculature to compensate for changes in perfusion pressure was unaffected by nerve section or nerve block and was designated 'autoregulation'. A similar phenomenon occurred in the liver (Ginsburg & Grayson, 1954), but the power of the liver to adapt its vascular resistance to changing perfusion pressure was destroyed by cannulating afferent or efferent vessels and was greatly reduced in the debilitated animal. There was, thus, evidence that factors outside the organ were also concerned and that the term 'autoregulation' was to some extent a misnomer. The term 'intrinsic regulation' is perhaps little better but was used because the regulation of blood flow is not strictly autonomous, although the focal point of this kind of blood-flow control is probably in the organ itself. The present experiments gave clear evidence that 'intrinsic regulation' also occurs in the brain. When the systemic blood pressure fell from 15 to 6 mm Hg little change occurred in cerebral blood flow although, during this period, cerebral vascular resistance progressively declined. Below 6 mm Hg vascular resistance continued to diminish but blood flow began to fall slightly. The fall in flow only became considerable, however, when the blood pressure dropped below 3 mm Hg. At pressures below 2 mm Hg it was usually impossible to detect any flow. In so far as there was still a positive perfusion pressure it must be concluded that below 2 mm Hg vascular resistance suddenly increased, possibly due to spasm of the vessels. The general blood-flow reactions of the brain to changing blood pressure were thus similar to those of the kidney and of the liver. They were similar, too, in that ganglion blocking agents had no effect on those reactions. Although it is hard to assess the precise action of these agents under conditions of hypotension, the conclusion is reasonable that nervous mechanisms involving extracerebral ganglia are not concerned in 'intrinsic regulation'. In other details the blood-flow reactions of the cerebrum to changing blood pressure differed from those of the liver. Thus, in the brain, far bigger falls in blood pressure occurred before there was any serious impairment of blood flow. Even with blood pressures as low as 3 mm Hg blood flow was diminished by less than 5%. This point may prove of importance in relation to haemorrhage and shock reactions; it is of immediate interest in view of the use of induced hypotension in clinical practice. A further point of difference lies in the terminal spasm of the cerebral vessels. This phenomenon might be explicable on the basis of critical closing pressure-the intravascular pressure below which any elastic vessel, by virtue of its physical structure, goes into spasm (Burton, 1952). In the liver, where

20 CEREBRAL BLOOD FLOW 29 much of the blood arrives by way of the portal vein, there is no evidence of terminal spasm and blood flow persists even with blood pressures of less than 1 mm Hg. A point of considerable importance in interpreting the results of other workers is that in our work cannulation of the jugular veins or other interference with the blood supply of the brain frequently abolished 'intrinsic regulation', and blood flow subsequently varied directly with the systemic blood pressure. It was also often difficult to demonstrate 'intrinsic regulation' after periods of anoxia, or anaesthetic overdose. We have no evidence as to the nature of 'intrinsic regulation' of intracerebral blood flow. If the experiments of Forbes & Wolff (1928) concerning the dilatation of pial vessels in response to lowered blood pressure are to be regarded as 'intrinsic regulation', then their observation that local application of cocaine did not affect the response may go even further to eliminate a nervous mechanism. Rosomoff & Holaday (1954) suggested that a critical controlling factor in the cerebral circulation might be the arterio-venous 2 difference, but whether this can be invoked in explanation of 'intrinsic regulation' remains to be seen. SUMMARY 1. An improved photographic method is described for the investigation of blood flow by 'internal.calorimetry'. Thermal conductivity was shown to be a function of blood flow in the cerebrum. 2. Blood-flow records from the frontal lobes of sheep, goats, cats and rabbits showed no spontaneous variations in the undisturbed, lightly anaesthetized animal. 3. Resting blood-flow levels were unaffected by ganglion blocking agents (tetraethylammonium chloride and hexamethonium bromide) or atropine. 4. Intra-arterial adrenaline caused cerebral vasoconstriction. Intravenous adrenaline increased blood flow-partly through the mechanical effect of the raised blood pressure and partly through a reflex vasodilator mechanism which was substantially or wholly abolished by the administration of atropine. 5. The evidence suggested that, in response to falling blood pressure, a fall occurred in intracerebral vascular resistance as a result of which blood flow in the frontal lobes was maintained relatively constant with blood pressure levels higher than about 4 mm Hg. When blood pressure dropped below 4 mm Hg there was a decline in blood flow. When the blood pressure fell below 2 mm Hg results suggested a sudden increase in cerebral resistance as an effect of which flow subsided to zero despite a positive perfusion pressure. 6. The preservation of relatively constant blood flow despite changing blood pressure (intrinsic regulation) is unaffected by ganglion blocking agents and is considered to be independent of external nervous control.

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