The density and nerve varicosity-smooth muscle cell separation of rabbit cerebral and ear arterial beds were compared. The rabbit middle cerebral artery and three of its successive branches and a comparable-sized ear artery and two branches were perfusion-fixed for electron microscopy and analyzed by quantitative morphometric procedures. The purpose was to determine if there are structural correlates to previously observed differences in the sympathetic control of these two vascular systems. The in vitro contractile response of isolated artery segments to electrical field stimulation of their intramural nerves is considerably less in cerebral arteries compared with the similar-sized ear arteries. Furthermore, in the cerebral but not the ear circulation, there is progressive diminution of the neurogenic response with successive branching. Although the total varicosity densities of the major ear and brain arteries studied are similar, and this parameter stays fairly constant with successive branching of the ear, it falls off considerably in the cerebral vessels. There is a significant difference in densities between the two vascular beds when "bare" varicosities located <1 pm from the medial smooth muscle are compared. The second-order branch of the ear artery has an average of 18 bare varicosities per 500-,m Tnhe effectiveness of sympathetic control of different parts of the vasculature shows remarkable variation, and some of the factors responsible for this have been identified. These include innervation density and the extent of separation of the innervation from the closest smooth muscle cells,1-3 transmitter receptor number,4 and adrenergic receptor affinity.5 Among the various vascular beds that receive a sympathetic innervation, perhaps the least responsive to sympathetic activity in vivo is the cerebral bed,6-8 and this has led to debate about its role, if any, in cerebrovascular regulation. In vitro studies show that isolated vessels from the rabbit cerebral bed respond, after a longer latency, more slowly and to a smaller extent than corresponding-sized arteries from the ear bed of the same species.9 Furthermore, in the cerebral circulation there is a progressive diminution of response in successive branches not only to sympathetic nerve activation Received February 22, 1993; accepted August 4, 1994. From the Department of Pharmacology, College of Medicine, University of Vermont, Burlington. Correspondence to John A. Bevan, MD, Department of Pharmacology, College of Medicine, Given Bldg, University of Vermont, Burlington, VT 05405-0068. © 1994 American Heart Association, Inc. circumference, and the corresponding cerebral vessel has only 2.8 bare varicosities per 500-,um circumference. The mean bare varicosity-smooth muscle cell separation (mean+SEM) is significantly (P<.05) less in the ear (1.18+0.06 ,um) than in the cerebral arteries (4.95 +0.23 jum). This is true of all vessels studied. Fifty-nine percent of the bare varicosities in the ear arteries are <1 ,um from the smooth muscle cells, and 1.2% are more distant than 5 gm. These values for cerebral vessels are 9.5% and 37%, respectively. In the ear vessels, 25% of the bare varicosities make close neuromuscular contact (within 500 nm of the smooth muscle), whereas only 3% do so in cerebral vessels; in cerebral compared with ear vessels, the percentage becomes significantly less with branching. These structural features of brain vessels, taken together with the lower sensitivity to and the diminished capacity to respond to norepinephrine, probably account for their weak neurogenic control. The results indicate that the cerebral circulation of the rabbit receives a sympathetic innervation that is relatively ineffective in altering cerebrovascular tone. (Circ Res. 1994;75:916-925.)

We studied isolated basilar artery segments from a rabbit model of chronic cerebrovasospasm. Autologous blood placed around the basilar artery of rabbits killed 1, 2, 3, 4, 5, 6, 7, or 9 days later caused narrowing of the segments with a biphasic time course. The first (immediate) phase was reversed by intra-arterial papaverine; the second phase exhibited an increasing component of narrowing that was papaverine-insensitive. Based on the passive force/length curves, basilar artery segments became increasingly stiff over 9 days. By contrast, the segments' contractility decreased. Responses of the basilar artery segments were greater over the first few days, but then became less than that of saline-injected controls. Contractions in response to norepinephrine and potassium were reduced. Endothelium-based acetylcholine-induced vasodilation progressively diminished, as did the response to sympathetic nerve stimulation. There was a negative correlation between artery wall stiffness and contractility. The papaverineinsensitive component of angiographic narrowing correlated directly with loss of contractility and with artery wall stiffness. These results are consistent with the conclusion that increased artery wall stiffness is a primary determining factor in the arterial narrowing of chronic cerebrovasospasm. {Stroke 1990;21:1478-1484)

OBJECTIVE : The dimensions and maximum wall tension capacity of segments of human pial arteries from normotensive obese and non-obese patients were compared. DESIGN : Segment size was assessed by quantitative morphometry of fixed sections and wall force by in vitro myography. SUBJECTS : Twenty normotensive non-obese and 13 normotensive obese humans body mass index (BMI) 22.4±0.5 and 33.9±1.7 kg/m 2 , respectively) were studied. RESULTS : There was no significant difference in the perimeter of the outer medial border, the smoothed out length of the internal elastic lamina, the ratios of media thickness to area and lumen diameter and the maximum wall force development between the two groups. CONCLUSION : Obesity per se is not associated with initial dimensional changes nor capacity to develop wall tension that might lead to the emergence of hypertension.

Pial arteries play a key role in the regulation of human cerebral blood flow. However, many of the features and mechanisms that regulate the tone and diameters of these vessels cannot be studied in situ. One approach is to study in vitro segments of arteries obtained during neurosurgical procedures. The ratios of arterial media thickness to lumen diameter and of the capacity to develop wall force to lumen diameter have important functional consequences and are known to change in disease. Experiments were carried out on pial arteries from normotensive humans to determine the way in which these parameters vary with vessel size. Vessel dimensions — media thickness and lumen diameter were derived from fixed sections using quantitative morphometry. Wall force was measured using a resistance artery myograph. The ratio of media thickness to lumen diameter and of maximum tension developed to lumen diameter both increased as vessel diameter decreased. These ratios do not change over the age range of 15–75 years. These findings show that although in vivo intralumenal pressure falls as human pial arteries become smaller, their media thickness and capacity to develop tone increase.

Responses of segments of basilar and middle cerebral arteries of eight human infants to activation of perivascular nerves and to vasoactive drugs were studied using a resistance artery myograph. The infants ages ranged from 23 wk of gestation to 34 postnatal days. Neurogenic vasoconstriction occurred in all segments and at 8 Hz was 12.7 ± 3.5% (11%) of tissue maximum and was blocked by phentolamine (10-6 M). There was no evidence of a neurogenic dilator response. Catecholamine histofluorescence was seen in nerves in the adventitia at all ages studied. Norepinephrine ED50 was 7.6 ± 1.8 × 10-7 M, and its maximum effect was 43.1 ± 5.7% of tissue maximum. Both neural and norepinephrine responses were greater than those of the proximal parts of adult human middle cerebral arteries obtained postmortem and surgically removed adult human pial arteries. Electron microscopy demonstrated that neural density at the adventitiomedial junction in the infant vessels was greater than in the pial arteries. Constrictor responses to serotonin and prostaglandin F2α were minimal in the two infants of 23 and 24 wk of gestation but were clearly present in the older infants. Histamine and acetylcholine were potent vasodilators. Indomethacin potentiated agonist-induced contraction. In a limited number of trials angiotensin II, neuropeptide Y, caused contraction and bradykinin, relaxation. It is concluded that there is a quantitative similarity between the studied responses of infant cerebral artery segments and human pial arteries of similar diameter. However, sympathetic nerves may potentially play a more important role in the regulation of cerebrovascular tone in the infant compared with the adult, and during the gestational period examined these vessels possess an indomethacin-sensitive system that buffers agonist tone.

BACKGROUND AND PURPOSE The primary goal of these studies was to understand and investigate the capacity of perivascular nerves to influence the tone of human pial arteries and to compare them with other human cephalic arteries, the superficial temporal and middle meningeal. METHODS Responses to electrical activation of intramural nerves and related features of fresh segments of human cephalic arteries-the pial (PA; 478+/-34 microm ID), middle meningeal (MMA; 540+/-41 microm ID), and superficial temporal (STA; 639+/-49 microm ID)-obtained from patients aged 15 to 82 years during surgical procedures were studied on a resistance artery myograph. RESULTS The PA segment responses to electrical nerve activation and to norepinephrine (NE; 10[-5] mol/L) were 1% and 21% of tissue maximum, respectively, compared with 6% and 34% for the MMA and 14% and 90% for the STA. Tissue maximum was defined as the force increase to 127 mmol/L KCl plus arginine vasopressin (1 microm). All arteries dilated well to acetylcholine. Possible explanations for the PA marginal neurogenic responses were assessed. NE ED50 was 5.4+/-2.2 X 10(-7) mol/L and did not vary with age or diameter. NE responsiveness did not increase in vessels with spontaneous or raised potassium-induced tone. Relaxation to isoproterenol was variable and propranolol did not increase the neurogenic response. Neither N(G)-monomethyl-L-arginine, N(G)-nitro-L-arginine methyl ester, endothelium removal, nor indomethacin consistently influenced the contractions to NE or neurogenic reactivity. The weak PA neurogenic response is in keeping with its poor innervation. As determined by catecholamine histofluorescence, innervation in the PA is sparse, with density increasing in the order PA, MMA, and STA. The incidence of nerve structures in the PA adventitio-medial junction was only 3% of those in the STA, and these were situated more than 3 microm from the closest smooth muscle cell. CONCLUSIONS We conclude that the weak neurogenic response of adult human pial artery reflects its poor innervation and responsiveness to NE, implying that these features are not important in the regulation of its diameter.

Segments of basilar and middle cerebral arteries of eight human preterm and early postnatal infants have been examined using the resistance artery myograph technique for wire-mounted segments and the pressure perfusion arteriograph. Myograph-mounted segments spontaneously developed tone of varying duration and time course. Perfused segments showed maintained tone levels of approximately 40% of maximum possible constriction when the intraluminal pressure was 60 mm Hg. This level is not different from that found in adult human pial arteries of similar lumen diameter. Indomethacin (10[-5] M) either initiated tone increase or potentiated existing tone in the isometrically mounted segments. After washout of vasoconstrictors norepinephrine (10[-6] M) and angiotensin II (10[-8] M), indomethacin caused a pronounced, long lasting increase in basal tone. Spontaneous tone was reversed by acetylcholine (10[-6] M), isoproterenol (10[-8] to 10[-5] M), histamine (10[-8] to 10[-5] M), and papaverine (10[-5] M). Low levels of tone were increased and higher levels decreased by intraluminal flow. The pressure/diameter curves of these vessels were of similar shape as those of the equivalent size in the adult. It is concluded that intrinsic tone is a prominent feature of these large cerebral arteries, and it is modified by an endogenous indomethacin-sensitive process.

BACKGROUND AND PURPOSE:Responses to changes in intraluminal pressure of isolated human pial arteries (200 to 1200 microns i.d.) obtained from patients undergoing neurosurgery were measured. METHODS:The vessels were cannulated and pressurized (60 mm Hg); vascular diameter and intraluminal pressure were recorded simultaneously. After spontaneous development of steady state tone, intraluminal pressure was changed to both higher and lower levels in random sequence. RESULTS:Human pial arteries exhibited myogenic responses and maintained their diameter over the pressure range of 20 to 100 mm Hg. The level of myogenic tone observed at 30 mm Hg did not vary significantly with artery diameter. In contrast, at 60 and 90 mm Hg, the extent of myogenic tone increased as the diameter decreased (up to 70% to 80% of maximum in 200-microns i.d. arteries). The arteries contracted to KCl 30 mmol/L, norepinephrine 1 mumol/L, and vasopressin 0.1 mumol/L and relaxed to acetylcholine 3 mumol/L. The extent of these responses did not vary with the diameter of the artery. Arterial distensibility, represented by the slope of the tangent of the passive pressure-diameter curve at lower pressures (5 to 50 mm Hg), increased as arteries became smaller. This is consistent with the possibility that the level of myogenic tone is related to vessel distensibility. Human omental arteries of comparable size did not develop myogenic tone but contracted to KCl and norepinephrine and relaxed to acetylcholine to an extent similar to pial arteries. CONCLUSIONS:There is a specific gradient of myogenic responsiveness in human pial arteries that varies inversely with their diameter. This tone does not develop in all vascular beds. These levels of tone in the pial circulation would be expected to be of profound functional significance by allowing blood flow to vary widely.

Human pial arteries obtained during surgery frequently exhibit spontaneous periodic contractions. Simultaneous measurements of membrane potential and vessel wall force were used to examine whether these contractions are associated with electrical activity of smooth muscle cells (SMCs). A total of 53 segments from 38 patients were studied, and of these, 26 showed spontaneous contractions related to periodic depolarization and generation of action potentials (APs). The resting membrane potential during the silent periods was -44.0 +/- 0.5 mV. APs without "overshoot'' were observed when spontaneous depolarization reached levels of -40 to -35 mV. Just over half of the arterial segments failed to exhibit spontaneous activity; however, APs could be generated during K+-induced depolarization. The mean SMC resting membrane potential of these vessels was -53.5 +/- 1.5 mV, and this value differed significantly from that of SMCs in spontaneously active arteries. Application of tetrodotoxin did not change the amplitude and duration of APs. Removal of Ca2+ from the bathing solution and addition of nifedipine completely inhibited the spontaneous APs and associated contractions. K+ depolarization failed to induce APs and contraction in the presence of nifedipine. We conclude that periodic spontaneous depolarization and AP generation underlie the periodic spontaneous contractions of human pial arteries. Both the APs and associated contractions are related to the activation of dihydropyridine-sensitive voltage-dependent Ca2+ channels. It is suggested that AP generation can be responsible for vasomotion of human pial arteries in vivo.

Smooth muscle cells (SMCs) in 58% of human pial arteries obtained during surgery showed no spontaneous contractions and displayed a stable resting membrane potential (MP) of -54.7 +/- 1.5 mV. Those that exhibited periodic spontaneous contractions associated with periodic depolarization and generation of spontaneous action potentials (APs) had a less negative MP of -43.1 +/- 0.5 mV (42%). Inhibition of calcium-activated potassium (KCa) channels in the silent arteries by charybdotoxin (CTX) and tetraethylammonium ions (TEA) induced dose-dependent depolarization, AP generation, and contraction. TEA and CTX enhanced the spontaneous depolarization and force in arteries that exhibited spontaneous activity. They also prolonged the spontaneous APs up to several times and increased their upstroke amplitude. Both TEA and CTX failed to produce significant depolarization in arteries treated with nifedipine. It is concluded that KCa channels are important regulators of human pial artery SMC resting MP and tone. They are also involved in the control of AP amplitude and duration and the associated contractions. These data suggest that alterations in the activity of SMC KCa channels could be responsible for the appearance of spontaneous activity in human pial arteries in vitro and that impaired function of these channels might be related to vasospastic phenomena in human cerebral circulation.

Cerebral arteriovenous malformations (AVMs) are vascular abnormalities that result in an excessive shunt of blood from the arterial to the venous system. Although there is brain parenchyma between the vascular channels of an AVM, it is often scarred and abnormal. The purpose of this study is to measure the elasticity of feeding vessels from AVMs obtained at the time of surgical resection. Pressure measurements of feeding arteries and draining veins of AVMs made prior to surgical resection and reported in the literature are reviewed. By combining the results of the in vivo experiments with published observations of pressure in vivo, a theory of how AVM hemodynamics can change over time is propose

In vitro studies of neural influences on cerebral arteries have for the most part been carried out on segments of the basilar artery, the Circle of Willis, or the origins of the major cerebral arteries (for example, refs. 1-4). In both animals and humans, constriction on electrical activation of intramural nerves has been observed, whereas dilation has been demonstrated only in nonhuman species. The relative size of neurogenic dilator and constrictor effects, and the variety of putative neurotransmitters and mechanisms, are remarkably different between species.4–6

The density and nerve varicosity-smooth muscle cell separation of rabbit cerebral and ear arterial beds were compared. The rabbit middle cerebral artery and three of its successive branches and a comparable-sized ear artery and two branches were perfusion-fixed for electron microscopy and analyzed by quantitative morphometric procedures. The purpose was to determine if there are structural correlates to previously observed differences in the sympathetic control of these two vascular systems. The in vitro contractile response of isolated artery segments to electrical field stimulation of their intramural nerves is considerably less in cerebral arteries compared with the similar-sized ear arteries. Furthermore, in the cerebral but not the ear circulation, there is progressive diminution of the neurogenic response with successive branching. Although the total varicosity densities of the major ear and brain arteries studied are similar, and this parameter stays fairly constant with successive branching of the ear, it falls off considerably in the cerebral vessels. There is a significant difference in densities between the two vascular beds when "bare" varicosities located < 1 micron from the medial smooth muscle are compared. The second-order branch of the ear artery has an average of 18 bare varicosities per 500-micron circumference, and the corresponding cerebral vessel has only 2.8 bare varicosities per 500-micron circumference. The mean bare varicosity-smooth muscle cell separation (mean +/- SEM) is significantly (P < .05) less in the ear (1.18 +/- 0.06 microns) than in the cerebral arteries (4.95 +/- 0.23 microns). This is true of all vessels studied. Fifty-nine percent of the bare varicosities in the ear arteries are < 1 micron from the smooth muscle cells, and 1.2% are more distant than 5 microns. These values for cerebral vessels are 9.5% and 37%, respectively. In the ear vessels, 25% of the bare varicosities make close neuromuscular contact (within 500 nm of the smooth muscle), whereas only 3% do so in cerebral vessels; in cerebral compared with ear vessels, the percentage becomes significantly less with branching. These structural features of brain vessels, taken together with the lower sensitivity to and the diminished capacity to respond to norepinephrine, probably account for their weak neurogenic control. The results indicate that the cerebral circulation of the rabbit receives a sympathetic innervation that is relatively ineffective in altering cerebrovascular tone.