Salvador Lluch

Development and Evaluation of an Experimental Model for the Study of the Cerebral Circulation in the Unanesthetized Goat

An animal model has been developed for the continuous measurement of total cerebral blood flow in the unanesthetized, unrestrained goat. We selected the goat because each internal maxillary artery, a branch of the external carotid artery, provides the total blood flow to each hemisphere via the rete mirabile. After the occlusion of the extracerebral vessels of the internal maxillary artery with thrombin, an electromagnetic flow transducer was chronically implanted on this artery, distal to the temporal artery, to measure hemispheric blood flow. Reproducible measurements of cerebral blood flow were obtained in ten unanesthetized goats for periods ranging between two weeks and five months. The mean cerebral blood flow for the ten goats was 133 ± 5 ml/min/100 gm tissue at an average mean aortic pressure of 91 ± 3 mm Hg, heart rate of 79 ± 3 beats/min, arterial PCO2 of 31.4 ± 1.0 mm Hg, P02 of 76.6 ± 1.7 mm Hg, and pH of 7.48 ± 0.02. The present experimental preparation allows cerebrovascular hemodynamics to be evaluated under near-normal conditions and is suitable for physiological and pharmacological studies in normal and abnormal states.

Pharmacological receptors of the cerebral arteries of the goat.

5-Hydroxytryptamine (5-HT), norepinephrine (NE), histamine (H) and potassium (K+) chloride induce dose-dependent changes in tension of the isolated middle crerbral artery of the goat. Vasopressin produces highly variable responses followed by tachyphylaxis; angiotensin II is ineffective over a wide dose range. The order of potencies of these vasoactive agents is 5-HT greater than NE greater than H greater than K+. With regard to their ability to induce maximal contractile responses, the order is: H greater than 5-HT, K+ greater than NE. Lysergic acid diethylamide (LSD) antagonizes the actions of 5-HT in a manner which progresses from surmountability to unsurmountability of the blockade depending on the concentration of LSD. The blockade exerted by LSD is reversed by washing. Phentolamine and diphenhydramine competitively antagonize the actions of NE and H, respectively. The potency of phentolamine and diphenhydramine in the cerebral arteries of the goat is similar to that determined in different tissues obtained from a variety of animal species. It is concluded that the cerebral arteries of the goat possess receptors for biogenic amines, the most effective of which is 5-HT; receptors for vasoactive peptides are ill defined.?25

Evidence for the Direct Effect of Adrenergic Drugs on the Cerebral Vascular Bed of the Unanesthetized Goat

Despite considerable research, the question of whether adrenergic drugs exert direct effects on the cerebral circulation has remained unresolved. With the development of a method for monitoring continuously the entire blood flow to one hemisphere in the unanesthetized goat, we have been able to study this problem directly. The effects of epinephrine, norepinephrine, and isoproterenol administered by close intra-arterial injection were investigated in 15 goats in which an electromagnetic flowmeter had been implanted previously on the internal maxillary artery, which, in this animal, provides the sole blood supply to a hemisphere. Both epinephrine and norepinephrine (0.1 to 5.0 μg) produced dose-dependent reductions in cerebral blood flow, a decrease of 55 ± 3% (SEM) occurring with the highest dose. Alpha receptor blockade of the ipsilateral hemisphere with phenoxybenzamine totally or partially abolished this cerebral vasoconstriction. Isoproterenol (0.01 to 1.0 μg) produced dose-dependent increases in cerebral blood flow, an increment of 75 ± 6% occurring with the highest dose. Beta blockade with propranolol totally or partially abolished the cerebral vasodilation induced by isoproterenol. Thus, epinephrine, norepinephrine, and isoproterenol exert powerful direct effects on the cerebral circulation of the unanesthetized goat, and these effects appear to be mediated by alpha and beta receptors.

Indirect adrenergic effect of histamine in cat cerebral arteries

SummaryHistamine (10(−4 M) induced an increase in the tritium outflow from cat cerebral arteries preloaded with 3H-noradrenaline. Pretreatment with reserpine (3 mg/kg, i.p., total dose) or removal of both superior cervical ganglia two weeks before the experiment abolished that increase. The presence of cocaine or diphenhydramine also prevented the rise in tritium efflux induced by histamine.Histamine (10(−8 M to 10(−3 M) elicited dose-dependent contractions in the isolated posterior communicating artery of the cat which were reduced in the presence of diphenhydramine at all doses except the highest three. The addition of phentolamine to the bath decreased the contractile responses at the doses lower than 10(−6 M. Pretreatment with reserpine or removal of both superior cervical ganglia also diminished the responses at doses of histamine below 10(−6 M and 10(−5 M, respectively. When cocaine was added to the bath there was a decrease in the contraction elicited at all doses except the last one.These results suggest the existence of an indirect adrenergic mechanism in the contractile response to histamine in cat cerebral arteries.

Adrenergic Mechanisms in Cerebral Blood Vessels: Effect of Tyramine on the Isolated Middle Cerebral Artery of the Goat

Tyramine induces dose-dependent changes in tension of the isolated middle cerebral artery of the goat. Cocaine, phentolamine and reserpine reduce the sensitivity of the tissue to tyramine by factors of 2.8, 3.7 and 3.7, respectively. The norepinephrine (NE) concentration of the arteries of the circle of Willis is 2.10 μg per gram and the corresponding value for the right atria is 1.25 μ per gram. Reserpine pretreatment (0.02 mg/kg/day for three days) reduces the NE concentration of the cerebral arteries to undetectable levels and that of the right atria to 2.4% of the control value. The relatively high concentration of NE of the cerebral arteries of the goat suggests that this tissue receives considerable sympathetic innervation. It is likely that part of the contractile response to tyramine is due to release of endogenous NE from sympathetic stores in the artery. However, some contractile response to tyramine remains after α-adrenergic blockade, reserpine pretreatment and in the presence of cocaine, suggesting that in addition to an indirect action (release of NE) tyramine also possesses a direct stimulatory effect in cerebral arteries.

Rete mirabile of goat: in vitro effects of adrenergic stimulation.

The carotid rete of the goat, a network of small arteries, is placed in the pathway of the main arteries which supply the brain. This structure lies within the cavernous sinus on each side of the pituitary. The presence of a carotid rete in many mammals has led to frequent speculations about its physiological function. The present study was designed to examine comparatively the responsiveness of goat retial and cerebral arteries to direct or indirect adrenergic stimulation. The contractile response of isolated retial arteries (150-500 micron in external diameter) to norepinephrine (10(-8) - 10(-4)M), field electrical stimulation (2-16 c/s), and tyramine (10(-6) - 10(-3) M) was negligible. In contrast, cerebral arteries of 300-500 micron in external diameter exhibited dose- or frequency-dependent contractions qualitatively similar to those previously described in larger cerebral arteries. The norepinephrine content in the walls of retial arteries was about 13% of that measured in cerebral arteries. These results suggest that the role of the carotid rete in the regulation of resistance to blood flow during adrenergic stimulation is negligible or nonexistent.

Analysis of cerebrovascular action of diazoxide in conscious goats.

The effects of diazoxide on cerebral blood flow were evaluated in unanesthetized goats under control conditions and after selective blockade of adrenergic or cholinergic receptors in cerebral vessels. Injections of diazoxide (1–27 mg) into the internal maxillary artery produced dose-dependent increases in cerebral blood flow1 an increase of 90% occurring with the highest dose. Administration of pnentolamine, propranolol, or atropine into the internal maxillary artery did not modify the cerebrovascular response to diazoxide. In reserpine-treated animals the cerebral effects of diazoxide were also unchanged. Intravenous Injections of diazoxide (150–400 mg) produced sustained hypotension and tachycardia whereas cerebral blood flow was maintained within normal values or increased slightly. The normal cerebral vasoconstriction obtained with injections of norepiUephrine directly into the Internal maxillary artery was unaffected during the diazoxide induced-hypotension. These findings snow that diazoxide exerts a powerful vasodllatory effect on cerebral vessels through mechanisms other than blockade of alpha-adrenergic receptors or inhibition of adrenergic activity. The results also indicate that activation of beta-adrenergic or atropine-sensitive vascular receptors in the cerebral response to diazoxide is negligible.

In vivo and In vitro studies on the cerebrovascular dilatation induced by diazoxide in normotensive and renal hypertensive goats.

We studied the in vivo and in vitro effects of diazoxide on the cerebral circulation of 8 normotensive (mean arterial pressure = 100 mm Hg) and 5 renal hypertensive (mean arterial pressure = 146 mm Hg) goats. Injections of diazoxide (03-27 mg) into the internal maxillary artery of unanesthetized goats produced dose-dependent increases in cerebral blood flow (electromagnetic flowmeter), this effect being significantly higher in hypertensive goats. Intravenous injection of 5 mg/kg of diazoxide into normotensive goats increased cerebral blood flow 40 ml/mln/100 g and mean arterial pressure dropped 22 mm Hg whereas in hypertensive goats cerebral blood flow was unchanged and mean arterial pressure decreased 50 mm Hg. The increase in heart rate due to intravenous diazoxide was similar in normotensive and hypertensive goats (35 beats/min). Cumulative applications of diazoxide (10-5 to 10-3M) on isolated middle cerebral arteries produced dilatory responses both under resting conditions and after previous tonic contraction by serotonin. This relaxation was significantly greater in arterial segments from hypertensive goats. The results indicate that diazoxide exerts powerful dilatatory effects on cerebral vessels, both in vivo and in vitro, and that these effects are particularly evident in hypertensive animals. Stroke, Vol 12, No 2, 1981

Relaxation of Isolated Middle Cerebral Artery Induced by Diazoxide

The effect of diazoxide on isometric tension of goat middle cerebral arteries was investigated both under resting conditions and under contraction produced by 10(-6) M serotonin. In addition, the inhibitory action of diazoxide was tested against contractile effects induced by different experimental interventions such as electrical field stimulation, norepinephrine, tyramine, histamine, and KCl. Diazoxide caused dose-dependent relaxation of cerebral arteries which was more pronounced when the vessels were previously contracted. High doses of diazoxide (10(-3) M) inhibited significantly the contraction induced by electrical field stimulation and all the vasoactive agents used. This inhibitory effect of diazoxide was greater for those drugs that directly or indirectly act through alpha-adrenergic receptor stimulation. Tritium release induced by electrical field stimulation of cerebral arteries previously labelled with 3H-norepinephrine was not affected in the presence of diazoxide. We conclude that diazoxide has a dilatory effect on goat brain vessels due to direct relaxation of smooth muscle together with a possible blockade of the alpha -adrenergic receptors. This effect might explain the maintenance of cerebral blood flow observed in vivo during diazoxide-induced arterial hypotension.