A BLOOD FLOW INCREASE TO A VISUAL STIMULUS IN THE OCCIPITAL LOBE OF THE CAT WITH BRAINSTEM TRANSECTION AT THE PRETRIGEMINAL LEVEL

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1 ACTA NEUROBIOL. EXP. 1977, 37: 5-14 A BLOOD FLOW INCREASE TO A VISUAL STIMULUS IN THE OCCIPITAL LOBE OF THE CAT WITH BRAINSTEM TRANSECTION AT THE PRETRIGEMINAL LEVEL K. SKOLASINSKA, L. KR~LICKI and B. ZERNICKI Isotope Laboratory, Institute of Physiological Sciences, School of Medicine and Department of Neurophysiology, Nencki Institute of Experimental Biology Warsaw, Poland Abstract. The effects of visual stimulation on the cerebral blood flow were studied in pretrigeminal cats. The visual stimulus was a feather duster which was moved for 4-5 min in.front of the cat's eyes. The cerebral blood flow was measured each 15 min with 133Xe clearance method in the left occipital lobe. The visual stimulation evoked an increase of the cerebral blood flow which was accompanied by an ocular following movements and a cortical EEG desynchronization. The arterial blood pressure was unchanged. All these responses habituated rapidly when the stimulus was repeated at 15 rnin intervals. A similar increase of the cerebral blood flow was obtained by g0/o C02 inhalation. The administration of amphetamine increased the cerebral blood flow, whereas Nembutal decreased it. The visual stimulus evokes an increase of the cerebral blood flow in a condition when the lower brainstem is separated from the cerebrum. INTRODUCTION Some recent data show that a visual stimulation can produce an increase of cerebral blood flow (CBF) in intact animals. Bondy et al. (3) reported the increase of the blood flow in the optic lobe of the chicken when it paid attention to the visual environment as shown by pecking the presented grain. Sokoloff's data (11) suggested changes in the blood

2 6 8. flow in the visual system of the restrained intact cat during repetitive flash stimulation. In the present work the CBF response to visual stimulation was investigated in cats with brainstem transected at the pretrigeminal level (1,13). In contrast to cerveau is016 preparation, the isolated cerebrum or" the pretrigeminal preparation is awake and works normally under many respects, which is shown by the presence of ocular orienting reflexes to visual stimuli, by a rapid habituation of these reflexes, and by the possibility of elaboration of conditioned ocular reflexes. The pretrigeminal cat is convenient for investigating the CBF response to visual stimulation for two reasons. First, the mechanism of this response is presumably simplified since it cannot be mediated by the lower brainstem which plays probably a role in the CBF regulation (7, 8). Second, in the pretrigeminal preparation the pain is eliminated and therefore the cat can be restrained without narcorsis in the stereotaxic apparatus for the purpose of adequate vislial stimulation and CBF recording. METHODS Eight cats were used. Under ether anesthesia a polyethylene catheter was inserted into the left lingual artery and the pretrigeminal brainstem transection was done at the rostropontine level (13). For the EEG recording two pairs of silver electrodes were placed on the dura mater in the right occipital and frontal areas. The interelectrode distance in each pair was 10 mm. For the EOG recording silver wire electrodes were placed subcutaneously above and below the right eyeball. The gammascintilator detector with a 4 cm NaJ crystal, dwelled 0.5 cm inside a 1 cm diam opening of a lead collimator, was positioned 1 cm above the left occipital lobe. The experiment was started after recovery of the cat from the posttransectional coma, as shown by the appearance of the ocular following reflex, usually about 2 h after the brainstem transection. The experiment lasted from 2 to 10 h and consisted of a number of successive 15 min trials (measurements). For measuring the cerebral blood flow, in the beginning of a trial a single bolus (0.05 ml) of 133Xe in physiological saline solution was injected through the lingual artery into the left external carotid artery and washout curve was recorded by ratemeter. The formula for the calculation of the CBF from the washout curve was based on Kety's approach for blood tissue exchange of inert gasses (6). The washout curve was plotted on the semilogarithirnic paper and divided in two compartments by graphical analysis (4). The half-times (T ) of both compartments were calculated from the respective slops (Fig. 1).

3 Using two different 1 (partition coefficient for the indicator between tissue and 'blood) for grey and white matter, the CBF for two compartments was calculated by the formula X I Flow = TI,, and expressed in ml/min/100 g of tissue. Con Fig. 1. An increase of the blood flow in the occipital lobe evoked by the movemeni of a feather duster. The records of two successive trials are presented. In the control trial (Con) the cat's eyes were closed, whereas in the visual trial (Vis) they were opened during the first 4 min and the visual stimulus was {presented (heavy line). Ordinate, counts per minute. The ce~ebral blood flow was calculated with two compartmental method and is expressed for grey matter (Fg) and white matter (F,) in ml/min/100 g. Cat. 11. During experiment the cat7s eyes were usually occluded. They were only opened during the first 4 or 5 min of the trials when a visual stimulus was applied (visual trials) and of some control trials. The visual stimulus was a feather duster moved by hand up and down with a speed of about 10 /s against a white screen located 50 cm from the cat's eyes. This stimulus evokes strong ocular and EEG responses in the pretrigeminal cat (9). All cats respirated spontaneously. The averaged C02 in the expiratory air varied in each preparation from 3 to 4 vol O/o. The arterial blood pressure, measured from the femoral artery, ranged in different preparations from 100 to 140 mm Hg. The body temperature was maintained at 38' For the description of the ECoG activity the terminology described previously (13) was used. Type I: almost continous low-voltage activity;

4 type 11: high-voltage activity mixed in different proportions with lowvoltage activity; type 111: high-voltage activity (mainly spindles) mixed with a small amount of low-voltage activity; type IV: high-voltage activity with delta waves occurring almost continually. These types of ECoG activity correspond loosely to the ECoG patterns present in the intact cat during the states of alert wakefulness, drowsiness, light synchronized sleep and deep synchronized sleep, respectively. RESULTS The effects of visual stimulation on the cerebral blood flow are shown in Fig. 1 and Table I. During the first trial with a feather duster presentation, the cerebral blood flow was clearly increased in both grey matter (CBFg) and white matter (CBFw) (t, = 2.74 for CBFg, t, = 2.75 for CBFw, P < 0.05, Student's t-test for correlated data). The average in- The effect of a repetitive visual stimulation on the blood flow in the left occipital lobe. Thr cerebral blood flow is expressed in ml/min/100 g and calculated separately for, grey mattee (CBFg) and white matter (CBFw). For each cat the data for a series of successive 15 min trials are presented (for cat 4, two series are shown). During control trials (Cor;) the cat's eyes were occluded. During the experimental trials (Vis) the eyes were opened during the first 4-5 min and the visuctl stimulus (feather duster) was presented. 1 CBFg 1 CBFw Cat / Con / ~on 11 4 after 2 h I Vis 1 I V I 2 ~ I Vis 3 I Con I Con I Con I Vis I I "is 2 1 Vis 3 1 Con crease was stronger for the grey matter. Two cats (cat 12 and 6) showing no increase of CBFg need comments. In cat 12 the CBFg increased in visual trial I1 in which the ocular following reflex was stronger than in visual trial I (Table 11). In cat 6 the ECoG activity flattened in the post-visual control trial and the cat died thereafter. Except the cat 10, the visual stimulus evoked a good ocular following reflex and in five cats it produced a clear desynchronization of ECoG activity (Table 11). In the mean arterial blood pressure and C02 level in the expiratory air no changes were observed.

5 The ECoG activity and ocular following reflex during repetitive visual stimulation. Data presented refer to the same cats and trials as in Table I. ECoG activity: type I, almost contirluous low-voltage activity, type 11, high-voltage activity mixed in different proportions with low-voltage activity; type 111, high-voltage activity (mainly spindles) mixed with a small amount of low-voltage activity. Following reflex: + + +, presumably adequate ocular following reflex during the whole trial; + +, originally adequate but rapidly habituating reflex; +, abortive reflex; -, lack of reflex. In cat 6 the ECoG activity flattened in the last control trial and the cat died after 10 min. Cat 4 after 2 h 5 I0 9 Predominant ECoG type Vis 1 1 Vis 2 1 Vis 3-1 Con I1 I 11 and 11' I I I and I1 / 11 I1 1 I I I I and I1 I I1 I I I ' 11 / I1 I Flattening Following reflex Vis 1 / Vis 2 ( Vis 3 In some control trials the eyes were opened but the feather duster was not presented. During these trials no increase of CBF was observed. In four cats the visual stimulus was presented during three successive trials (Table I and 11). In cats 11, 4 and 5, where the CBF increased during the first visual trial, it returned almost to the control value during the second and third visual trials. An mixed design analysis of variance for these three cats showed differences among the set of trials consisting of three visual trials and the preceding control trial (F,,, = 9.31, F3,g;,.,, = 6.99, P < 0.01, for CBFg; F,,, = 5.12, F,,,;,,,, = 3.86, P < 0.05 for CBWw). For the grey matter, the Duncan test showed differences on the level P < 0.01 between a visual trial I and other trials, and no differences among other trials. For the white matter, the visual trial I differed at the level P < 0.05 from the control trial and the visual trial 11, but there were no differences between the control trial and visual trials I1 and 111. The ocular following reflex and the ECoG desynchronization seemed to 'habituate less rapidly than the CBF response (Tables I and 11). In three cats type of ECoG activity changed spontaneously during an experiment. In these cats a higher blood flow in the grey matter was present during type I ECoG activity than during the types I1 and I11 (Table 111). In the white matter this effect was unclear.

6 Dependence of the blood flow in the occipital lobe on the spontaneous changes of type of ECoG activity. Mean data are presented (n = number of observations). CBFg 1 Type I n 1 Type I1 n I CBFw Type I11 n 1 Type 1 Type I1 1 Type UI Three cats were given a small dose of Nembutal (10 mgl'body weight, i. v.). In two cats it was administrated about the end of experiment. In all cases the ECoG activity became more synchronized and the cerebral blood flow (both in the grey and white matter) decreased moderately (Table IV). Mean arterial blood pressure and CO, level in the expiratory air did not change. Decrease of the blood flow in the occipital lobe after Nembutal injection. The data for two successive trials are presented. CBFg CBFw I ECoG type Before I After Before ( After To four cats amphetamine (0.5-1 mgjkg body weight, i.v.) was administrated in the time when the following reflex was poor or absent (in three cats in the early period of experiment). Then in all cats appeared the following reflex and the ECoG activity became more desynchronized in three cases (Table V). In the grey matter the CBF increased considerably (t, = 4.79, P < 0.05, Student's t-test for correlated data). In the white matter the effect was not observed. The presentation of the visual stimulus did not produce a further increase of the CBF (Table V). The mean arterial blood pressure increased in all preparations, by 40 mm Hg on the average. About the end of experiment the effect of an increased C02 level in the inspiratory air was examined in three cats. For this purpose the cats were paralized with Tricuran and artificially ventilated. They were

7 Increase of the blood flow in the occipital lobe after amphetamine injection. The data for three successive trials are presented. In the third trial (Vis) a feather duster was presented under amphetamine action. I CBFg I ECoG type - Cat Before IAfter 1 Vis 1 Befor After ( Vis ~eforel~fter I Vis - CBFw given 9010 CO, for about 10 min. The measuring of cerebral blood flow started when the ECoG activity became clearly more desynchronized (Table VI). At that time the COP in the expiratory air was about vol O/o. The CBF increased in the grey matter (t, = 6.43, P < 0.01, Student's t-test for correlated data). In three cats the increase of CBF in the white matter was observed; this effect, however, was statistically insignificant. The mean arterial blood pressure increased on the average by about 10 mm Hg. TABLE VI Increase of the blood flow in the occipital lobe evoked by 9% C02 inhalation. The data for three successive trials (before, during and after C02 inhalation) are presented. The increase of the CBF wasaccompanied by ECoG desynchronization. In cat 12 the 9% C02 was applied two times. CBFw ECoG type Cat / ~eforel~uringl After ~eforel~uring After ~eforel~uringl After I ' :I I11 : II , IV 1111andIY / after 2 h / / In cats 5, 9 and 6 the episodes of ECoG flattening were observed (in cat 6 such episode preceded its death). During ECoG flattening the CBF in both grey and white matter remained at the control level (similar observation was reported by Ingvar and Soderberg, 5). However, neither the visual stimulus nor C02 application could at that time evoke any increase of the CBF.

8 DISCUSSION Our experiments showed that a strong visual stimulus (the movement of a feather duster) caused a sharp increase of the blood flow in the occipital lobe of the pretrigeminal cats. The mean size of the blood flow response was comparable to that produced by an increased level of C02 in the inspiratory air. Thus the reactivity of cerebral circulation to visual stimulation seems to be virtually intact in the pretrigeminal cat. Some authors (5, 12) described an increase of the CBF in cats during electrical stimulation of several points in the pons and midbrain. In our preparations the transection was just behind the pontine-midbrain junction, and thus during visual stimulation the possible way of the CBF increase mediated by the lower brainstem and the truncus sympathicus was eliminated. In addition, the arterial blood pressure obviously could not be increased by visual stimulation. We can conclude that the lower brainstem is not critical for the CBF increase evoked by visual stimulus. A higher value of CBF was often associated with ECoG desynchronization. The CBF increase evoked by the visual stimulus was usually associated with ECoG desynchronization. However, with repetition of visual trials, the CBF response seemed to habituate more rapidly than the ECoG response. A spontaneously appearing type I ECoG activity was accompanied by an increase of CBF, which however was clearly smaller, than during visual stimulation. Finally, small doses of Nembutal produced both a synchronization of the ECoG activity and a decrease of the CBF. These data are in accordance with the observations of other authors (2, 11). An amphetamine administration evoked a desynchronization of the ECoG activity and a strong increase of the CBF. However, the mechanism of amphetamine action was certainly more complex. It affected also the lower brainstem of our preparations, as shown by an increase of the arterial blood pressure. Results of Bondy et al. (3) on chickens suggest that the attention of the animal to a presented visual stimulus is critical for CBF increase. In our cats a strong CBF increase was always associated with an excellent ocular following reflex, but not vice versa, as shown by the probable more rapid habituation of the CBF response. In summary we conclude that (i) the CBF in the pretrigeminal cat (and probably also in the intact cat) is correlated with the level of the animal's arousal (measured by ECoG desynchronization), and (ii) during visual stimulation the CBF is correlated with a specific attention to this stimulus (measured by the intensity of the following reflex). However, there is probably other factor (or factors) which contribute to the value

9 of CBF. The CBF increase may be correlated, for example, with the intensity of fear produced by the feather duster movements. Drives certainly can be present in the isolated cerebrum of the pretrigeminal cat as shown by the possibility of rapid elaboration of classical and instrumental ocular conditioned reflexes in this preparations (13). Our observations on the CBF were restricted to the occipital lobe and we do not know to what extent they can be refered to the whole brain. However, it is reasonable to think that in the occipital lobe the effect of the visual stimulus was the strongest. In the pretrigeminal preparation the ECoG arousal to the visual stimulus is clearly strongest in the occipital cortex (13). Sokoloff (11) demonstrated on cats that CBF is affected by visual stimulation mostly in the visual system. Finally, Risberg and Ingvar (10) showed regional changes in the CBF during mental activity. Recently it was found in this Laboratory (9) that the same visual stimulus (feather duster movement) produced a dramatic change in the hippocampal EEG activity of the pretrigeminal cat. It would interesting to know whether it was associated with local increase of CBF in the hippocampus, which might contribute to the CBF response observed in our preparations. We thank Professor M. Purves, Professor Stella Niemierko and Dr P. Jastreboff for comments and Mrs. Janina Rokicka, Mrs. Jagoda Michalska and Mr. J. Folga for technical assistance. This work was supported by Projects and 432iVI of the Polish Academy of Sciences. REFERENCES 1. BATINI, C., MORUZII, G., PALESTINI, M., ROSSI, G. F. and ZANCHETTI, A Effects of complete pontine transection on the sleep-wakefulness rhythm: the miqpontine pretrigeminal preparation. Arch. Ital. Biol. 97: BLADY-MOULINIER, M. and INGVAR, D. H EEG frequency content related to regional blood flow of cerebral cortex in cat. Exp. Brain Res. 5: BONDY, S. C., LEHMAN, R. A. W. and PURDY, J. L Visual attention affects braip blood flow. Nature 248: H0EDT-RESMUSSEN, K., SVEINSDOTTIR, E. and LASSEN, N. A Regional cerebral blood flow in man determined by intraarterial injection of radioactive inert gas. Circulation Res. 18: INGVAR, D. H. and SODERBERG, U Cortical blood flow related to EEG patterns evoked by stimulation pf the brain stem. Acta Physiol. Scand. 42: KETY, S. S The themy and applications of the exchange of inert gas at the lungs and tissues. Pharmacal. Rev. 3: MEYER, J. S., TERAURA, T., SAKAMOTO, D. and KONDO, A Central neurogenic c6ntrol of cerebral blood flow. Neurology 21:

10 8. MOLNAR, L. and SZANTO, J The effect of electrical stimulation of the bulbar vasomotor centre on the cerebral blood flow. Quart. J. Exp. Physiol. 49: RADIL-WEISS, T., ZERNICKI, B. and MICHALSKI, A Hiapocampal theta activity in the acute pretrigeminal cat. Acta Neurobiol. Exp. 36: RISBERG, J. and INGVAR, D. H Regional changes in cerebral blood volume during mental activity. Exp. Brain Res. 5: SOKOLOFF, L Local cerebral circulation at rest and during altered cerebral activity induced by anesthesia or visual stimulation. In S. S. Kety and J. Elkes (ed.), Regional neurochemistry. Pergamon Press, Oxford, p SUZUKI, H. and TUKAHARA, Y Cerebral circulation during arousal reaction of EEG. Tohoku J. Exp. Med. 84: ZERNICKI, B Isolated cerebrum of the pretrigeminal cat. Arch. Ital. Biol. 112: Accepted 14 October 1976 K. SKOLASINSKA and L. KROLICKI, Institute of Physiological Sciences, School of Medicine, Krakowskie Przedmiehcie 26/28, Warsaw, Poland. B. BERNICKI, Nencki Institute of Experimental Biology, Pasteura 3, Warsaw, Poland.