J. Physiol. (I956) I33,

Transcription

1 232 J. Physiol. (I956) I33, A STUDY OF THE EFFECT OF THE PATTERN OF ELECTRICAL STIMULATION OF THE AORTIC NERVE ON THE REFLEX DEPRESSOR RESPONSES By W. W. DOUGLAS, J. M. RITCHIE AND W. SCHAUMANN* From the National Institute for Medical Research, Mill Hill, London, N. W. 7 (Received 19 April 1956) The systemic blood pressure of an animal is controlled by reflexes involving the barosensory fibres in the carotid sinus and aortic nerves and their associated receptors. The adequate stimulus to the baroreceptors produces in these fibres not a discharge of a steady continuous nature, but bursts of impulses synchronized with the fluctuations in systemic blood pressure which accompany the cardiac cycle: and it has been suggested that this grouping of the afferent impulses might be of importance for the central mechanisms involved in the buffer reflexes (Bronk & Stella, 1932; Ead, Green & Neil, 1952). We have therefore studied the effect of interrupted pulsatile electrical stimulation of the aortic nerve and compared it with the effect produced by stimulation with a continuous train of shocks. METHODS The experiments were done on adult lop-eared rabbits anaesthetized by urethane g/kg given as a 25% solution into the marginal ear vein. The vagi, aortic nerves and sympathetic trunks were cut low in the neck. One aortic nerve was dissected free of the surrounding tissue and placed on a pair of Ag-AgCl electrodes for stimulation. In some experiments the action potentials set up in the nerve were recorded monophasicallv through a pair of platinum electrodes placed near the cut end, amplified in a directly coupled amplifier and displayed on a cathode-ray tube. The dissected region was flooded with liquid paraffin held in a pool by arrangement of the skin flaps. Records of systemic blood pressure were taken from a femoral artery and registered with a mercury manometer on a kymograph. Both common carotid arteries were ligated so as functionally to denervate the carotid sinuses. The interrupted, pulsatile, electrical stimulation was achieved in the following way. A master oscillator produced a train of triggering pips at a given frequency (Fig. 1 a). A voltage pulse of variable width (Fig. 1 b) operated an electronic gate allowing only the pips shown in Fig. 1 c to pass. These pips were further gated by a rectangular wave form (Fig. 1 d) of variable width and repetition rate. The surviving pips (Fig. 1 e) triggered a conventional electronic stimulator with an RF output unit (Schmitt, 1948) attached to it. In this way a pulsatile stimulus pattern similar to that shown in Fig. 1 e was produced. The total period of stimulation, the duration of the bursts and the interval between them, and the frequency of the shocks within the burst were all independently * British Council Scholar.

2 STIMULUS PATTERN AND DEPRESSOR REFLEXES 233 variable. The number of shocks delivered to the nerve during any given period of stimulation was counted by means of a dekatron scaler unit. The width of each shock was usually 1OO,usec. The intensity of the stimulus used was such as to give maximal stimulation of the medullated fibres in the nerve but was too weak to excite the non-medullated depressor fibres of unknown origin which are also present in this nerve and have been recently described by Douglas, Ritchie & Schaumann (1956). a Fig. 1. b c d e. liii liii liii The method of producing an interrupted train of stimulating shocks. For explanation see text. RESULTS Prolonged periods of continuous trains and interrupted trains of stimuli In these experiments the aortic nerve was stimulated until the blood pressure ceased to fall and stimulation then continued for a further 2-3 min, the pressure being thus kept at the lowered level. During this period the pattern of stimulation was changed so that the effect of a continuous train of impulses at a steady frequency could be compared with that of an interrupted or pulsatile train of impulses. It was arranged that the total number of shocks delivered during a given period of interrupted stimulation was the same as that delivered during an equal period of continuous stimulation. The method used to do this is illustrated diagrammatically in Fig. 2. In this example, the frequency of application of shocks during the continuous type of stimulation was 16/sec (Fig. 2a). During the interrupted stimulation, the stimulator frequency was raised to 32/sec or to 64/sec but, as shown in Fig. 2 b, c, the shocks were applied to the nerve only during one-half or one-quarter of the time respectively. In this way the total number of shocks applied during any given stimulation period was kept constant for the three types of stimulation and the average frequency of stimulation kept at 16 shocks/sec. Experiments of this sort were done with average frequencies of 8, 16, 32 and 64 shocks/sec. The number of bursts delivered per minute during the interrupted stimulation was varied in order to mimic various heart rates. The bursts were given 240 times/min to mimic a normal heart rate, 360 times/min to mimic a fast heart rate and 60 or 120 times/min to mimic a slow heart rate. At all these rates essentially similar results were obtained. At 8 shocks/sec there was no appreciable effect on blood pressure whether the stimulus was continuous or interrupted. At low and moderate average frequencies of stimulation (i.e shocks/sec) continuous and interrupted

3 234 W. W. DOUGLAS, J. M. RITCHIE AND W. SCHAUMANN stimulation were usually equally effective whether the duration of each burst in the interrupted stimulus was equal to a half or a quarter of the interval between successive bursts. Typical results are shown in Figs. 3 and 4. These figures also illustrate that the results obtained were similar whether the frequency of stimulation was such as to produce strong responses (Fig. 3) or weak responses (Fig. 4). At 64 shocks/sec the effects of continuous and interrupted stimulation were again the same when the duration of each burst was 16: I a 32: 1lii liii liii liii liii liii b c 1 sec Fig. 2. An example of three different types of stimulation with the same average frequency, and the explanation of the cipher used to describe them briefly. In the three traces a, b and c, the total number of impulses delivered in a second is 16. In a they are evenly spaced and this type of stimulation is described by the cipher 16:1. In b and c the same number of stimuli (i.e. 16 in a sec) are crowded together in bursts (240 bursts/min), each of which lasts for a half and a quarter respectively of the interval between bursts and the instantaneous frequencies during the burst are 32 and 64 shocks/sec respectively. The types of stimulation illustrated in b and c are described symbolically as 32: J and 64: i respectively, i.e. the left-hand number in the cipher gives the stimulator frequency and the right-hand fraction gives the duration of the burst. equal to half the interval between successive bursts (middle trace, Fig. 3) but not when the burst lasted only a quarter ofthis interval (i.e. when the frequency of shocks within each burst was high, shocks/sec), the effect of the interrupted stimulus being markedly smaller (Fig. 5). In some experiments, however, particularly those in which low frequencies of stimulation were used, interrupted stimulation was more effective than continuous stimulation. This effect was never very marked and the biggest differences of this kind which we observed are illustrated in Fig. 6. Short periods of continuous and interrupted stimulation Experiments were done to compare the effects of continuous and interrupted trains of stimuli using periods of stimulation of 4 sec. The 4 sec period was sufficiently long for substantial depressor effects to be produced and yet short enough to allow a large number of observations to be made in a comparatively short time. During each period the pattern of stimulation was kept fixed: after successive periods the frequency of the shocks during the bursts and the duration of the burst were changed but in such a way as to keep constant the total number of shocks in the 4 sec period, i.e. the pattern of stimulation was

4 STIMULUS PATTERN AND DEPRESSOR REFLEXES 235 Fig. 3. The depressor response to stimulation of the central end of an aortic nerve of a rabbit under urethane anaesthesia. In each record, during the periods marked by the solid horizontal bars the shocks were delivered in a continuous train. During the periods marked by the broken horizontal bars, the same number of shocks was delivered in a given time but in groups 240 times/min (the average frequency of stimulation for each record is thus kept constant). The cipher below each bar represents the stimulator frequency and the duration of the burst. Fig. 4. The depressor response to stimulation of the central end of an aortic nerve of a rabbit under urethane anaesthesia. In each record, during the periods marked by the solid horizontal bars the shocks were delivered in a continuous train. During the periods marked by the broken horizontal bars, the same number of shocks was delivered in a given time but in groups 240 times/min, top and bottom record, and 360 times/min, middle record. (The average frequency of stimulation for each record is thus kept constant.) The cipher below each bar represents the stimulator frequency and the duration of the burst.

5 236 W. W. DOUGLAS, J. M. RITCHIE AND W. SCHAUMANN changed but the average frequency of stimulation was kept constant. Thus in a typical experiment depressor responses were obtained using stimuli applied at rates of 64 and 128 shocks/sec delivered in bursts lasting for i and i respectively of the interval between bursts. The average frequency in each of these two tests was thus 32 shocks/sec. For comparison a third response was obtained using a steady frequency of 32 shocks/sec. Fig. 5. The depressor response to stimulation of the central end of an aortic nerve of a rabbit under urethane anaesthesia. During the periods marked by the solid horizontal bars the shocks were delivered in a continuous train. During the periods marked by the broken horizontal bars, the same number of shocks was delivered in a given time but in groups 240 timet/min (the average frequency of stimulation is thus kept constant). The cipher below each bar represents the stimulator frequency and the duration of the burst. Fig. 6. The depressor response to stimulation of the central end of an aortic nerve of a rabbit under urethane anaesthesia. In each record, during the periods marked by the solid horizontal bars, the shocks were delivered in a continuous train. During the periods marked by the broken horizontal bars, the same number of shocks was delivered in a given time but in groups 240 times/min (the average frequency of stimulation for each record is thus kept constant). The cipher below each bar represents the stimulator frequency and the duration of the burst.

6 STIMULUS PATTERN AND DEPRESSOR REFLEXES 237 In the first group of experiments the bursts during the interrupted type of stimulation were delivered at a rate of 240 bursts/min, and such experiments were made using average frequencies of 16, 32 and 64 shocks/sec. The results obtained were similar to those which have been described above for prolonged stimulation. The response to interrupted stimulation was the same as that to continuous stimulation when the average frequency was 32 shocks/sec and the 140-9I Fig. 7. The depressor response to 4 sec periods of stimulation of the central end of an aortic nerve of a rabbit under urethane anaesthesia. In each of the three records there are three depressor responses: the two outermost responses were obtained with stimulation at a steady frequency and the central response was obtained with the same average frequency but with the shocks delivered in bursts each of which lasted for a quarter of the interval between the bursts. In record a the average frequency of stimulation was 32 shocks/sec and the bursts were applied at a rate of 240/min. In record b the average frequency of stimulation was 64 shocks/sec and the bursts were applied at a rate of 240/min. In record c the average frequency of stimulation was 32 shocks/sec and the bursts were applied at a rate of 60/min. The horizontal line represents 1 min. frequency of stimuli within each burst was 64 or 128/sec, and was also the same when the average frequency was 64 shocks/sec and the frequency of stimuli within each burst was 128 shocks/sec (Fig. 7a). The response to interrupted stimulation was, however, less when the average frequency was 64 shocks/sec but the instantaneous frequency of the shocks within each burst was high,

7 238 W. W. DOUGLAS, J. M. RITCHIE AND W. SCHAUMANN 256 shocks/sec (Fig. 7b). When the average frequency was 16 shocks/sec very small depressor responses were obtained. In such tests interrupted stimulation produced similar or greater effects. In the second group of experiments the bursts of stimuli were applied more frequently (360/min) or less frequently (60/min). When the bursts were delivered at a rate of 360/min the results obtained were like those which have been described for a rate of 240/min, but when the bursts were delivered at a rate of 60/min the interrupted stimulation was usually less effective than continuous stimulation (Fig. 7c). A typical experiment is illustrated by Fig. 8. _20 _ VI2 0 10~~~1 0-0~~~~~~~~~ Duration of bursts (sec) 1 m Fig. 8 Fig. 9 Fig. 8. The relationship between the depressor response and the burst duration. The ordinates are the peaks of the depressor responses produced by 4 sec periods of stimulation of a rabbit's aortic nerve with a continuous tramn of shocks and with trains where the same numbers of shocks were delivered in bursts of different durations: the abscissae are the burst durations. The bursts occurred 60 times/min. The average frequency was 32 shocks/sec. Fig. 9. Depressor responses to stimulation of the central end of an aortic nerve of a rabbit under urethane anaesthesia. The responses are those to a 4 sec period of stimulation followed by a prolonged period of stimulation. The frequency of stimulation is indicated by the number above each record. The average frequency stimulation was 32/sec and depressor responses were obtained using stimuli applied at rates of 64, 128 and 256 shocks/sec delivered every second in bursts lasting for 2, i and i sec: a fourth response was obtained to a steady frequency of stimulation of 32 shocks/sec. These responses have been plotted against the duration of the bursts (Fig. 8), and it is clear that the magnitude of the response (when the bursts were applied 60 times a minute) was not wholly independent of the pattern of stimulation and that the reflex response became greater the more evenly the shocks were spread over the stimulation period. Similar elresut were obtained when the average frequency of stimulation was 64 shocks/sec but with a low average frequency of 16 shocks/sec the response was usually independent of the stimulus pattern.

8 STIMULUS PATTERN AND DEPRESSOR REFLEXES 239 Frequency response curves with long and short periods of stimulation The finding that in experiments using bursts of stimuli at a rate of 60/min the pattern of stimulation did not influence the response to a long period of stimulation in the same way as it did that to a 4 sec period of stimulation led us to examine whether or not differences were to be found in the frequency response curves determined using the two stimulation periods. The procedure adopted was to stimulate the aortic nerve for a period of 4 sec, wait until the depressor response was waning, and then stimulate until a maintained response occurred (Fig. 9). This procedure was repeated at different frequencies of L /V -o r- ~~0 0 L 0~ ~~ ~ L / 0. 1 Teq v~~~ ~Fg 10 0ig I 00 E0, d! Frequency of stimulation (shocks/sec) Frequency of stimulatfon (shocks/sec) Fig. 10 Fig..11 Fig. 10. The relationship between the depressor response and frequency of stimulation of the central end of an aortic nerve of a rabbit under urethane anaesthesia. Open circles: with long periods of stimulation. Closed circles: with 4 sec periods of stimulation. Fig. 11. Same experiment as in Fig. 10. Each point is the ratio of the response obtained with the 4 sec period of stimulation to that obtained with prolonged stimulation. stimulation and the sizes of the falls occurring with long and short periods of stimulation were plotted against frequency (Fig. 10). The general shape of both frequency/response curves was the same: both showed a maximal response at 128 shocks/sec, and fell off rapidly with frequency to be about 20% of this value at 16 shocks/sec. At all but the lowest frequency used the ratio of the two responses remained almost constant tending to rise slightly with frequency (Fig. 11). In another experiment the influence of the duration of the stimulus upon the curve relating the frequency of stimulation to the reflex depressor response was

9 240 W. W. DOUGLAS, J. M. RITCHIE AND W. SCHAUMANN determined using periods of stimulation lasting for 1, 2, 4, 8 and 16 sec. The maximum response for any particular period of stimulation occurred when a frequency of 128 shocks/sec was used. In Table 1 the responses obtained when frequencies of 32 and 64 shocks/sec were used are expressed as a percentage of the corresponding maximal responses obtained using 128 shocks/sec. For both frequencies this percentage was scarcely affected by the duration of stimulation. These experiments thus show that much the same frequency/response curve is obtained whether the stimulus be applied for long periods so that maintained responses are obtained, or for short periods (1-16 sec) when only transient, smaller responses are obtained. TABLE 1. Effect of electrical stimulation of the aortic nerve on arterial blood pressure Response as percentage of Period of response at 128 shocks/sec stimulation,a_ (sec) 32 shocks/sec 64 shocks/sec DISCUSSION Bronk & Stella (1932), after providing an elegant demonstration of pulsatile and non-pulsatile discharges in single barosensory fibres in the carotid sinus nerve in different conditions, raised the question whether or not the grouping of the afferent impulses into discrete volleys, which is the normal pattern of afferent activity in this nerve, may be an important factor in regulating the activity of the centres upon which they impinge. Evidence does exist, from experiments in which the reflex effects of pulsatile and non-pulsatile pressure changes in the carotid sinus have been compared, that a pulsatile pressure of the same mean value as a steady pressure is the more effective in eliciting the typical responses to barosensory excitation. For example, McCrea & Wiggers (1933) came to the conclusion that variations in pulse pressure rather than changes in mean pressure dominated the production of reflex cardiac changes produced by the baroreceptors ofthe carotid sinus, and Strauss (1940) has shown that the reflex hypotension induced by increased pressure in the carotid sinus was better maintained by pulsatile pressure and that the 'blutdruckcharakteristik' obtained with pulsatile pressure differed from that obtained with steady pressure, in that a pulsatile pressure produced a greater reflex hypotension than did a steady pressure of the same mean value. Recently, Ead et at. (1952) found that when a steady flow through the innervated sinus region was converted to a pulsatile flow, the blood pressure fell and remained at its lower level. They also recorded the electrical activity in

10 STIMULUS PATTERN AND DEPRESSOR REFLEXES 241 some of the sinus nerve barosensory fibres and observed that changing from a steady to a pulsatile flow in the sinus caused a grouping of the previously continuous discharge in the fibres into bursts of impulses at a higher frequency, but that the total number of impulses occurring in a given time sometimes remained unchanged. They concluded that the grouped impulses were more capable of affecting the vasomotor centre than a steady discharge of an equal number of impulses at a lower frequency; and that the greater depressor effect caused by the pulsatile flow was the result of this grouping and was a reflex response to an alteration in the pattern of the impulse activity in the sinus barosensory fibres. Our technique has allowed us to test more directly the influence of the pattern of the buffer nerve discharge on the response of the vasomotor mechanisms, and the results obtained-that we have been unable to produce changes anything like as large as those illustrated by Ead et al. (1952), who found that a change from non-pulsatile to pulsatile perfusion of one sinus could cause a fall of 50 mm Hg-lead us to question their conclusion that the grouping of impulses plays an important part in the greater effectiveness of the pulsatile pressure stimulus. The smallness of our effect cannot be attributed to the intervention of any known buffering systems; for in our experiments the vagi and aortic nerves were cut and the carotid sinus regions were presumably functionally denervated by the ligation of both common carotid arteries. Our results in fact show that within wide limits the vasomotor response to electrical stimulation of all the fibres of the aortic nerve depends mainly on the number of shocks applied in a given time and hardly at all on the pattern in which they are applied. Ead et al. (1952) were led to their conclusion largely by the finding that their records of electrical activity in a few fibres (usually the larger fibres) showed no great change in the total number of impulses discharged during pulsatile and non-pulsatile pressure changes in the sinus; and from the extrapolation that the total number of impulses from all the fibres to the vasomotor centre was similar in the two conditions. This argument takes no account of the role played by additional fibres which are recruited when a steady pressure in the sinus is converted to a pulsatile one and which seems to us to be the more likely explanation of the greater efficacy of pulsatile stimulation. The records obtained by Ead et al. (1952) at low perfusion pressures do in fact show that a large fibre quiescent at steady pressure becomes active when the pressure is made pulsatile as is to be expected, not only because of the higher instantaneous pressures reached during pulsation, but also because, as shown by Bronk & Stella (1932), the efficacy of the stimulus is determined by the rate of change of pressure as well as by the mean pressure attained. The possibility of such recruitment certainly exists even at fairly high intra-sinusal pressures because there are some barosensory fibres which are not activated until pressures of up to 150 mm Hg are reached (Landgren, 1952). 16 PHYSIO. CXXXIII

11 242 W. W. DOUGLAS, J. M. RITCHIE AND W. SCHAUMANN SUMMARY 1. A comparison has been made of the depressor effects of stimulation of the rabbit's aortic nerve with continuous and interrupted trains of electrical shocks, in order to find out whether or not the grouping of impulses or pattern of stimulation influences the response. 2. When stimulation is prolonged sufficiently to cause a maintained depressor response this response is usually independent of the pattern of stimulation. Only at low rates of stimulation is the interrupted type of stimulus slightly more effective. 3. These results suggest that any greater depressor effect of pulsatile pressure changes in the great vessels over a steady pressure, is not attributable to the different pattern of impulses set up by the pulsations. It is suggested that the most important factor involved is the recruitment of fibres. 4. When stimulation lasts for only 4 sec and causes only a transient depressor response, interrupted stimulation may be more, equally or less effective than a non-pulsatile stimulus, depending on the rate at which bursts are delivered and on the average frequency of stimulation. 5. The relationship between the depressor responses and the frequency of stimulation remains substantially the same whether determined with brief periods of stimulation (2-16 sec) producing transient responses or with longlasting stimulation producing maintained responses. REFERENCES BRONK, D. W. & STELLA, G. (1932). Afferent impulses in the carotid sinus nerve. J. cell. comp. Physiol. 1, DOUGLAS, W. W., RITCHIE, J. M. & SCHAUMANN, W. (1956). Depressor reflexes from medullated and non-medullated fibres in the rabbit's aortic nerve. J. Phy8iol. 132, EAD, H. W., GREEN, J. H. & NEIL, E. (1952). A comparison of the effects of pulsatile and nonpulsatile blood flow through the carotid sinus on the reflexogenic activity of the sinus baroceptors in the cat. J. Physiol. 118, LANDGREN, S. (1952). On the excitation mechanism of the carotid baroceptors. Acta physiol. 8cand. 26, MCCREA, F. D. & WIGGERS, C. J. (1933). Rhythmic arterial expansion as a factor in the control of heart rate. Amer. J. Physiol. 103, ScEMITT, 0. H. (1948). A radio frequency coupled tissue stimulator. Science, 107, 432. STRAUSS, E. (1940). Die Bedeutung der Druckamplitude und des Herz-Reflexes fur die reflektorische Selbststeuerung des Kreislaufes. Arch. Krei8lForsch. 6,