Disappearance of catecholamine fluorescence from the adrenergic nerves in arterial grafts in rats: an experimental fluorescence histochemical study.

The disappearance of catecholamine fluorescence from the noradrenaline-containing sympathetic nerve fibres after arterial transplantation was studied using a femoral artery graft sutured to rat carotid artery. Glyoxylic acid-induced fluorescence was used to demonstrate adrenergic nerves histochemically. At six hours the network of fibres had started to degenerate, and catecholamine fluorescence from the adrenergic nerves had almost completely disappeared within 24 hours of grafting. Control specimens from normal femoral arteries showed a dense network of fluorescent adrenergic nerves. Based on observations of the relatively rapid liberation of catecholamines from the degenerating adrenergic nerves, we suggest that catecholamines liberated from degenerating adrenergic nerves may have an important role in early vasospasm in microvascular and coronary bypass surgery.     

Degeneration and regrowth of adrenergic nerves after microvascular anastomosis. A fluorescence histochemical study on end-to-end anastomoses of femoral vessels in the rat

The normal femoral artery and its branches were found to be innervated with a dense network of adrenergic nerves. The nerve plexus around the vein was sparse. Adventitial stripping of the femoral vessels, with or without division and reanastomosis, caused local disappearance of catecholamine fluorescence in the stripped area. The distal adrenergic innervation remained normal if the femoral nerve was left intact. Division of the femoral nerve, alone or in combination with blood vessel division and reanastomosis, caused total disappearance of catecholamine fluorescence from the adrenergic nerves of the entire distal neurovascular tree examined. At the end of the observation period of 36 weeks from the time of division of the nerve, artery and vein with subsequent microvascular anastomosis, numerous adrenergic nerves were observed to have crossed the suture line. The vascular nerve plexus around the femoral vessels was dense in places, but in other places sparse or absent. It seems that the reinnervation occurs not only over the suture line, but also together with other regenerating nerves from the adjacent tissues and by collateral sprouting from adjacent adrenergically normally innervated areas.     

Histochemical study of adrenergic nerve fibers in the skeletal muscle of the rat]

The distribution of adrenergic nerve fibers in the vessels, nerves, and skeletal muscles of the rat was studied by the glyoxylic acid fluorescence method and by electron microscopy. The changes in catecholamine fluorescence were investigated after decentralization, lumbar sympathectomy, and femoral nerve ligation. Catecholamine fluorescence was mainly localized in the arteries, and arterioles in the perimysium of the skeletal muscle. Sometimes, a spot-shaped catecholamine was seen in the metarteriole and capillary. Adrenergic nerve fibers ran parallel with femoral nerve fibers. After lumbar sympathectomy catecholamine fluorescence disappeared, the fluorescence in the quadriceps femoris muscle did not disappear even after ligation of the femoral nerve. Unmyelinated nerve fibers were identified adjacent to the capillary, whereas adrenergic nerve fibers were seen approaching to the quadriceps femoris muscle within a gap of 300 nm.     

Innervation of syngeneic vein grafts in the rat. The regeneration of adrenergic nerves

Regeneration of adrenergic nerves was studied sequentially in segments of the supradiaphragmatic inferior vena cava transplanted from one rat into the abdominal aorta of another of the same inbred strain. Nontransplanted vein segments were examined as controls. The adrenergic nerves were demonstrated by using the glyoxylic acid-induced fluorescence method and the formaldehyde-induced fluorescence method for the demonstration of cathecholamines. A total of 16 syngeneic grafts were examined 3, 5, 8, 16, 27, and 32 weeks after transplantation. Three additional grafts were wrapped inside a silicone tube during the operation to prevent regeneration of nerves from the surroundings. These rats were then killed 8 weeks after the operation. In the control specimens the supradiaphragmatic inferior vena cava was innervated with a rather dense plexus of adrenergic nerves showing varicosities. No nerves were seen in the 3-week-old grafts. In the 5-week-old grafts some sparsely distributed solitary nerve fibres were seen. In the 8-week-old grafts sparsely distributed regenerated nerves reinnervating the vasa vasorum and nerve plexa were observed. The 16-, 27-, and 32-week-old grafts showed rather poorly innervated areas and areas where the nerve plexus was dense. One graft showed very dense reinnervation with morphologically abnormal adrenergic nerves forming “droplet fibers” and showing dense accumulations of catecholamines. The grafts surrounded with silicone tubes had no adrenergic nerves and showed only some regenerated nerves crossing the suture line. The results of the present study indicated that syngeneic inferior vena cava transplanted into the abdominal aorta will be reinnervated with adrenergic nerves. The pattern of the nerve regeneration remains incomplete and patchy.     

Plasma dopamine-beta-hydroxylase activity and catecholamine levels in anesthetized dogs following acute hemorrhage.

Because dopamine-beta-hydroxylase (DBH) is released from storage vesicles in adrenergic nerves and the adrenal medulla along with catecholamines, determination of circulating levels of this enzyme might serve as an index of sympathoadrenal activity. This hypothesis has been studied in dogs anesthetized with cyclopropane, isoflurane, and thiopental that were subjected to a single acute hemorrhage and followed for 5 hours. Plasma DBH activity and catecholamine levels were determined before and every 30 minutes after the hemorrhage. Changes in DBH activity did not correlate well with changes in levels of circulating catecholamines in the dog in response to this form of stress.     

Effect of distension on adrenergic innervation of the rat urinary bladder

Summary The effect of distension on adrenergic innervation was investigated in the rat urinary bladder. Bladders were distended for 3 h by forced diuresis and ballon obstruction, and specimens were taken from the bladder dome, body and neck for the demonstration of glyoxylic acid-induced fluorescence of catecholamines. Depletion of catecholamines started after 10 h and was almost complete after 2 days. The fluorescence had recovered part way after 5–7 days and was practically normal after 21 days. Small, intensely fluorescent (SIF) cells in the ganglia continued to leak catecholamines throughout the 21-day study period. The primary clinical success of distension therapy for the treatment of unstable bladder may be at least partly due to a reversible disturbance in the function of the adrenergic nerves, which have an excitatory alpha-adrenergic dominance in such cases, but the persistent leakage from SIF cells raises the question of whether distension causes prolonged disturbances in bladder function.     

Failure of perivascular sympathectomy to remove adrenergic nerves from peripheral vessels of the rabbit ear skin.

A perivascular sympathectomy was performed at the base of the ear artery in 11 New Zealand white rabbits. Two days later three were killed, and the central nerve was cut in the remaining eight. The contralateral ear served as a control. Specimens were taken from the distal parts of both ears two days later and the glyoxylic acid-induced fluorescence method was used to show the adrenergic nerves. The arteries of the normal, control ear were surrounded by a dense plexus of fluorescent adrenergic nerves, which were abolished by perivascular sympathectomy only in the segment from which the adventitia had been removed. The adrenergic innervation was normal proximal to the site of perivascular sympathectomy, but there was a short segment of the central vessel distally in which it was diminished. No changes in adrenergic innervation were found in the distal third of the rabbit ear.     

Increased spillover of norepinephrine to the portal vein during CO-pneumoperitoneum in pigs

BACKGROUND: Elevated intra abdominal pressure (IAP) during CO2-insufflation has been associated with increased catecholamine concentrations in plasma. We have previously indicated that this may be due to a regional increased spillover from the abdominal region. In this experimental study we investigated catecholamine spillover from the drainage area of the portal vein during CO2-pneumoperitoneum. Methods: Eight pigs under general anesthesia were investigated before and after CO2-pneumoperitoneum with an IAP of 15 mmHg. Regional spillover of catecholamines was determined by measuring plasma catecholamine concentrations and flow simultaneously. Plasma concentrations of catecholamines were measured from the portal and femoral veins, the pulmonary and carotid arteries. Flow data were collected with laser-Doppler transit time flow probes around the portal and femoral veins. Cardiac output was measured by the thermo-dilution technique. Estimated spillover was calculated by the veno-arterial difference multiplied by flow. RESULTS: We found a significant increase in estimated spillover of norepinephrine from the drainage area of the portal vein from 10 (-1.2, 78) ng x min(-1) to 27 (1.8, 475) ng x min(-1)[median (range)] (P = 0.05), but no change in estimated spillover of norepinephrine from the drainage area of the femoral vein. Plasma concentrations of norepinephrine increased in central venous and arterial blood. There was no significant change in epinephrine concentrations in arterial blood. CONCLUSION: Estimated norepinephrine spillover from the drainage area of the portal vein increased during CO2-pneumoperitoneum in pigs. This may indicate that the increased norepinephrine concentrations found in arterial plasma reflects a local activation of sympathetic nerves in the region of the portal drainage area.     

Myocardial catecholamines following cold cardioplegic arrest during open-heart surgery

Preservation of both right and left ventricular subendocardial and subepicardial muscle was assessed using quantitative polarization microscopy (birefringence measurements) with preservation of myocardial catecholamines measured by fluorescence microscopy in biopsies from 20 consecutive patients who underwent open heart surgery with cold cardioplegic arrest (St. Thomas' Solution). Six of the 7 patients with clinical complications were predicted from the birefringence results. One developed left ventricular deterioration during bypass, two patients right ventricular deterioration, one patient both left and right ventricular deterioration and two patients had poor left ventricular function before bypass. Birefringence measurements were thus reliable in predicting post-operative cardiac outcome. There were no significant changes during the bypass period in the catecholamine scores, even in those patients who had clinical complications. Fluorescence microscopy showed that the "free" myocardial nerve net and the pericoronary nerve plexuses retained their catecholamine stores equally well. This indicated that St. Thomas' cardioplegia preserves myocardial catecholamine stores, depletion of which would remove a potentially important compensatory mechanism in cardiac pump failure. There may however be a temporary blockade in the release of endogenous cardiac catecholamine (noradrenaline) stores from the adrenergic nerve terminals following cold cardioplegic arrest despite myocardial pump failure.     

An evaluation of resting arterial ischemia models in the rat hind limb

Techniques for using the rat hind limb as a model of pure arterial ischemia at rest have not been well defined. Because the rat has no profunda femoral artery, numerous collateral pathways exist to the hind limb, and femoral artery ligation is not an effective method of inducing arterial ischemia. After several anatomic studies, a two stage operation to produce arterial ischemia in the left hind limb was devised. The first stage involved surgical interruption of collateral and re-entrant vessels, and the second stage involved femoral artery ligation. Using Xenon 133 clearance as an estimate of blood flow, reduction in flow to 14, 24, and 37% of the simultaneously measured value in the right hind limb was obtained at 2 hours, 2 days, and 5 days post ligation. Oxygen extraction in the left hind limb doubled both at 2 hours and at 2 days post ligation. Histological evaluation of the anterior compartment musculature after 5 days demonstrated loss of nuclei, degenerating contractile elements, edema, and inflammatory infiltrate. Evaluation of rats that had undergone isolated femoral artery ligation showed a 66% reduction in flow 2 hours after ligation, but no reduction in flow at 5 days, no increase in oxygen extraction, and only nuclear changes on histological exam at five days.     

Adrenergic innervation of the striated muscle of the intrinsic external urethral sphincter from patients with lower motor spinal cord lesion

The adrenergic innervation of smooth and striated muscle components of the intrinsic external urethral sphincter from patients with suprasacral lesions and detrusor-sphincter dyssynergia has been described previously, when no adrenergic nerves were found associated with striated muscle fibers. In our study the intrinsic external urethral sphincter from patients with lower motor neuron lesions and detrusor areflexia was studied histochemically using the glyoxylic acid method to visualize catecholamines. Varicose adrenergic nerves were demonstrated in the smooth muscle. Adrenergic nerve fibers also were found along the edge of individual striated muscle fibers as well as around striated muscle bundles. Blood vessels in both regions of the urethral sphincter were innervated by adrenergic nerves. We conclude that in patients with lower motor neuron lesions and detrusor areflexia there is a substantial invasion by adrenergic nerve fibers in relation to smooth and striated muscle in the urethra, although the function of the nerve fibers is not known.     

Functional innervation patterns in the corpus spongiosum and glans in the dog

The neural control of smooth muscle cells in the corpus spongiosum, helicine artery and bulbus glandis of the dog was investigated in relation to the mechanism involved in erection, using isometric tension recording and micro-electrode methods. In the corpus spongiosum, field stimulation evoked twitch-like contractions followed by relaxations. These relaxations were enhanced and prolonged by neostigmine and partly suppressed by atropine. Guanethidine abolished the twitch-like contractions and increased muscle tone. The relaxations observed after pre-treatment with guanethidine were abolished by tetrodotoxin (TTX), thereby indicating that these muscles are innervated by adrenergic excitatory, cholinergic and non-adrenergic non-cholinergic inhibitory nerves. In the helicine artery and bulbus glandis, field stimulation evoked contractions and these contractions were abolished by guanethidine or TTX, indicating that these muscles are innervated by adrenergic excitatory nerve fibres. After pre-treatment with guanethidine and atropine, muscle relaxation appeared in response to field stimulation in the helicine artery but not in the bulbus glandis, indicating that the helicine artery in the corpus spongiosum is also innervated by non-adrenergic non-cholinergic inhibitory nerves in addition to the excitatory adrenergic nerves. In the smooth muscle cells of the corpus spongiosum, slow potential changes were correlated with spontaneous contractions and field stimulation evoked excitatory or inhibitory junction potentials. The neural mechanism involved in erection is discussed in relation to the topical difference in the autonomic innervation patterns in the corpus spongiosum, helicine artery and bulbus glandis.     

Colonic nerve network demonstrated by quinacrine

Quinacrine can be used for fluorescence microscopy to visualize certain cell bodies and fibres in the myenteric plexus of the intestine, forming a network different from the cholinergic and adrenergic systems. In the Piebald mouse, animal model of congenital megacolon, the quinacrine positive plexus is thin in the distal narrowed segment and well developed in the large proximal zone. In Hirschsprung's disease, the quniacrine positive network has the same pathologic features but, contrary to the animal model, there is an increased number and size of extrinsic cholinergic and adrenergic fibres. So this mouse is an animal model of a pseudo-Hirschsprung's disease, which has not yet been reported in human pathology.     

Haemodynamic effects of verapamil administration after large doses of fentanyl in man

Thirteen patients with good left ventricular function undergoing coronary artery revascularization were studied to determine the cardiovascular effects of verapamil, 75-150 micrograms X kg-1, after a large dose (100 micrograms X kg-1) of fentanyl, with pancuronium for muscle relaxation. The patients were continued on their usual cardiovascular medications until the time of surgery, which included nitrates, beta adrenergic blockers, and nifedipine. Anaesthesia with fentanyl was associated with decreases in mean arterial blood pressure, systemic vascular resistance, left ventricular stroke work index, and circulating catecholamine levels. Mean values were not further changed by verapamil, but individual patients had additional modest decreases in blood pressure and systemic vascular resistance. Cardiac index, however, was well maintained. Plasma catecholamines remained depressed after verapamil under the study condition. Thus, in patients with good left ventricular function, clinically relevant doses of verapamil were well tolerated even in the presence of an anaesthetic that included large doses of fentanyl, with suppression of circulating catecholamine levels.     

Vein to artery grafts. An experimental study of reinnervation of the graft wall.

Iliolumbar vein to iliac artery grafts were placed in 21 rats by microsurgical techniques. Graft innervation was examined at five time intervals between 1 and 32 weeks after surgery. Nerve fibers were demonstrated microscopically by formaldehyde-induced fluorescence of catecholamines. The morphology and degree of graft innervation were assessed, semiquantitatively, relative to the contralateral iliac artery (control) within each animal. Nerves were seen in the graft region as early as 2 weeks, but it was not until 4 weeks that they were present along its length (5 mm). The formation of a nerve plexus in the adventitia surrounding the graft was evident at 8 weeks. By 16 weeks the degree of innervation in the graft had increased to a level that was greater than the control iliac artery in three of four animals examined. Grafts at 32 weeks were also hyperinnervated. However, the morphology of this innervation was different from the control arteries; nerve fibers were finer, not varicosed, and were located at a greater distance from the outer layer of smooth muscle cells. The origin of the nerves appeared to be collateral sprouts from nerves supplying the adjacent iliac vein and also from invading vasa vasorum. The host iliac artery nerve plexus did not contribute to graft innervation.     

Adrenergic innervation and stimulation of canine urethra

Sympathetic nervous innervation of the canine urethra was studied using catecholamine histofluorescent staining and the in vitro muscle bath. Morphologically, adrenergic nerves were found terminating on the urethral smooth muscle. Pharmacologically, urethral strips responded like muscle strips from the bladder base to adrenergic stimulation. This evidence supports the concept that the sympathetic nervous system supplies and modulates the function of the smooth muscle of the urethra and that urethral changes noted with adrenergic stimulation are not secondary solely to stimulation of the rich periurethral vaculature nerve supply.     

Beta-adrenergic receptor changes during coronary artery bypass grafting

To evaluate whether the function of beta-adrenergic receptors, essential to the biologic activity of catecholamines, is altered during coronary artery bypass grafting, we measured, in 16 patients undergoing myocardial revascularization, the density and the affinity of lymphocyte beta-adrenergic receptors before anesthesia induction (control) and at the end of cardiopulmonary bypass. Variations in the density and affinity of beta-adrenergic receptors were determined in vitro. Repeated determinations of plasma epinephrine and norepinephrine concentrations were also performed. Overall, no significant modification was observed in mean density and affinity of beta-adrenergic receptors at the end of cardiopulmonary bypass when compared with control values. However, a significant decrease (p less than 0.05) in affinity for isoproterenol was found in the six patients who had high catecholamine levels during cardiopulmonary bypass. In contrast, no significant modification of beta-adrenoreceptor affinity for isoproterenol was observed in the 10 patients who did not have this degree of adrenergic activation. In addition, beta-adrenoreceptor affinity for isoproterenol was decreased in the three patients in whom intraaortic balloon pumping was mandatory after discontinuation of cardiopulmonary bypass. We suggest that this decreased affinity of lymphocyte beta-adrenergic receptors could be related, at least in part, to a sustained adrenergic activation occurring in some patients during cardiopulmonary bypass.     

Effects of transient ischemia on nutrient flow and arteriovenous shunting in canine hindlimb.

A canine model was developed to simulate use of a pneumatic tourniquet in the clinical setting in order to study the acute and delayed effects of transient ischemia on limb and tissue blood flow, using radioactive microspheres and electromagnetic flow probes. Experimental femoral artery flow rose markedly after tourniquet ischemia, and remained significantly elevated for 24 hours (p less than 0.01). Blood flow to the rectus femoris and anterior tibial muscles rose significantly (p less than 0.05) immediately after tourniquet ischemia, and the latter remained significantly elevated at 24 hours (p less than 0.05). Blood flow to the skin of the experimental limbs was elevated significantly (p less than 0.05), immediately ater tourniquet ischemia, and at no other time. Blood flow to the nerves did not increase to its maximum until 15 minutes after tourniquet deflation, and by 24 hours was normal. Tibial and femoral marrow blood flow remained significantly lower in the experimental limb throughout the 24-hour period. Ater tourniquet ischemia (300 mmhg; 2 hours), greatly increased femoral artery flow was related to reactive hyperemia in skin, muscle, and nerve. Twenty-four hours after tourniquet deflation, there was a significant increase in femoral artery and anterior tibial muscle blood flow, and significant arteriovenous shunting from the experimental limb. These data suggest that the hemodynamic response to a transient ischemic injury is more prolonged and of greater magnitude than previously believed.     

Effects of Hydrocortisone, Phenoxybenzamine and Propranolol on the Blood Plasma Levels of Adrenaline and Noradrenaline During Hemorrhagic Hypotension in the Dog

The adrenergic aspects of the mechanism(s) of the vasodilating action of phenoxybenzamine and massive doses of hydrocortisone were studied in anesthetized dogs subjected to controlled hemorrhagic shock. During the predrug hypotension period with decreased cardiac output and heart rate, the plasma adrenaline and noradrenaline levels were both increased. Phenoxybenzamine injected alone augmented the cardiac output and heart rate and reduced the plasma adrenaline without changing the plasma noradrenaline concentration.
However, if the adrenergic alpha-receptor blocker was administered in combination with massive doses of hydrocortisone, the degree of vasodilation increased further with a concomitant increase in the concentrations of the two plasma catecholamines. Thus there seems to he no clear-cut relationship between hemodynamic changes and plasma catecholamines. The present findings illustrate the extreme complexity of plasma catecholamine kinetics during hypovolemic shock.     

The anatomy of erection

The authors review the current knowledge concerning the anatomy of erection. The nervi erigentes pass through the hypogastric plexus, travel along the cavernous nerve and terminate in the smooth muscle of the erectile bodies. There appears to be three types of nerves: adrenergic nerves, cholinergic nerves and non-adrenergic, noncholinergic nerves. The arterial blood supply is derived from branches of the internal pudendal artery. The venous drainage occurs via the circumflex veins towards the deep dorsal vein of the penis. The corpus cavernosum is composed of a thick, extensible albuginea which contains an extensible vascular network surrounded by smooth muscle tissue, which, when it contracts, appears to induce detumescence and, when it relaxes, appears to allow inflow of blood into the erectile bodies.