The neural and humoral pathways in remote limb ischemic preconditioning.

Remote ischaemic preconditioning (RIPC) is a therapeutic intervention that has been demonstrated to reduce myocardial injury in the clinical setting. However, the underlying cardioprotective mechanisms remain unclear. We hypothesised that RIPC utilises both humoral and neural pathways to convey the cardioprotective signal from the preconditioned remote organ to the heart. C57BL/6 mice were anaesthetised and subjected to in vivo 30 min coronary artery ischaemia followed by 120 min of myocardial reperfusion, at the end of which myocardial infarct size was measured and expressed as a percentage of the risk zone. RIPC was induced by 3 cycles of 5 min left femoral artery occlusion interspersed with 5 min reperfusion before prolonged myocardial ischaemia with or without femoral vein occlusion (humoral pathway), femoral nerve resection and/or sciatic nerve resection (neural pathway). RIPC resulted in a smaller myocardial infarct size when compared to control. However, occluding the femoral vein completely abolished the infarct-limiting effect of RIPC. Similarly, combined femoral and sciatic nerve resection also abolished the cardioprotective effect of RIPC. Interestingly, femoral nerve or sciatic nerve resection alone only partially abolished the infarct-limiting effect of RIPC. In conclusion, remote limb ischaemic preconditioning reduced myocardial infarct size in the mice in a manner which implicates both a neural and humoral pathway.

[Baroreflexes originated in vertebral artery zones upon peripheral vein tonus, systemic arterial blood pressure, and external respiration].

The investigation was intended to study the role ofbaroreceptors ofhemodynamically isolated zone of vertebral arteries in regulation of peripheral veins tonus, arterial pressure and external respiration. Pressure decrease in this vascular reflexogenic zone led to reflex responses of increase in femoral vein tonus, elevation of blood pressure level and stimulation of external respiration. The opposite reflex responses of cardio-respiratory functional system to initial pressure activation of vertebral arteries baroreceptors are observed. Basing on generalization of our own findings and similar physiological and morphological researches of other authors, it is established that afferentation from the vertebral artery zone is a reflexogenic factor of somatic muscles' veins tonus regulation. These reflexes of capacity vessels tonic activity changes are part of cardio-respiratory responses of maintaining the tissue gaseous exchange.

Leg blood flow during slow head-down tilt with and without leg venous congestion.

The effects of slow changes in body position on leg blood flow (LBF) were studied in nine healthy male subjects. Using a tilt table, sitting volunteers were tilted about 60 degrees backwards to a supine position within 40 s. To modify the venous filling in the legs, the tilt manoeuvre was repeated with congestion of the leg veins induced by two thigh cuffs inflated to a subdiastolic pressure of 60 mmHg. Doppler measurements in the femoral artery were used to estimate LBF. Additional Doppler measurements at the aortic root in five of the subjects were taken for the determination of cardiac output. The LBF was influenced by body position. In the control experiment it increased from 500 ml x min(-1) in the upright to 780 ml x min(-1) after 15 min in the supine position. A mean maximal value of 950 ml x min(-1) was observed 20 s after the tilt. Heart rate remained almost constant during the tilt phase, whereas stroke volume increased from 90 ml to 120 ml and it remained at that level after the cessation of the tilt. Congestion of the leg veins had no significant effect on heart rate, stroke volume and mean blood pressure. However, it increased vascular resistance of the leg during and after the tilt. After 15 min in the tilted position LBF amounted to 600 ml x min(-1). The results suggest that the filling of the leg veins is inversely related to leg blood flow. The most likely mechanism underlying this observation is a local effect of venous filling on vasomotor tone.

The sympathetic efferent innervation of the cutaneous and muscle veins in cats. A comparative study using retrograde localization with horseradish peroxidase.

Previous studies using electrical stimulation, tension recording and fluorescence histochemical methods indicate that variations in the sympathoadrenergic innervation exist in regional veins of the limb. In the present experiment, we used horseradish peroxidase (HRP) as a retrograde tracer to localize and quantitate the sympathetic innervations to the saphenous, femoral and muscle veins in the cat. The animal was anesthetized with pentobarbital. In three groups of cats, a segment of saphenous (n = 8), femoral (n = 8) and muscle (n = 10) vein was isolated. HRP was applied on the outer vein wall for 3-4 h to allow uptake into the nerve endings. The paravertebral sympathetic chain on the same side of HRP application was dissected after the animal was killed and fixed 60 h following the application of tracer. HRP-labeled neurons were counted in each sympathetic ganglion from L1 to S1. The average number of neurons (mean +/- SE) were 913 +/- 99, 732 +/- 70 and 234 +/- 32, respectively, for saphenous, femoral and muscle vein. There was no statistical difference between the saphenous and femoral vein (P > 0.1). The muscle vein was far less innervated (P < 0.001). When the surface area (mm2) of the vein segment for HRP application was taken into account, the neurons per mm2 were 44.1 +/- 4.8 for saphenous vein, 24.6 +/- 1.8 for femoral vein and 10.2 +/- 1.3 for muscle vein. The neuron density was significantly different among the three groups (P < 0.01). In a single ganglion, the distribution of HRP-neurons appeared to be scattering in pattern.(ABSTRACT TRUNCATED AT 250 WORDS)

Localization of sympathetic postganglionic neurons innervating the femoral-saphenous vein in cats.

Physiological and histochemical studies have suggested that the limb veins are innervated by sympathetic adrenergic fibers. In the present experiment, horseradish peroxidase (HRP) was used as a retrograde tracer to identify and localize the sympathetic postganglionic neurons that innervate the femoral-saphenous vein in cats. In anesthetized cats, HRP was applied perivascularly on a femoral and a saphenous vein segment (4-8 mm in length for each segment) to allow uptake into the nerve endings. The sympathetic chains on both sides were dissected after the animal was sacrificed and fixed 60 h following the HRP application. Histological examination on serial section was done to count the HRP-labeled neurons in each sympathetic ganglion from L1 to S1. In 10 cats, the total number of HRP neurons amounted to 8569. Most neurons arose from L3 (47%) and then L4 (31%). The number of neurons became progressively decreasing towards both ends of the sympathetic chain. Few neurons (less than 2% of the total) were discovered in the contralateral sympathetic ganglia. In each ganglion, the distribution of HRP neurons appeared to be scattering. Our findings provide anatomical evidence to support that the femoral-saphenous vein of the cat was innervated by the sympathetic efferent fibers. The main origins of these neurons are the third and fourth lumbar sympathetic ganglia.

Peptide-containing nerves in the rat femoral artery and vein. An immunocytochemical and vasomotor study.

Peptide-containing nerves have been examined in the rat femoral artery and vein using immunocytochemical and vasomotor techniques. The general neuronal marker PGP 9.5 revealed a moderate supply of nerve fibres and fascicles forming a loose network in the adventitia and the adventitial-medial border of the artery and vein. The majority of the nerve fibres in both the artery and vein displayed immunoreactivity for neuropeptide Y (NPY) and tyrosine hydroxylase (TH). The distribution pattern and number of these two types correlated well. The artery had a slightly richer PGP 9.5- immunoreactive nerve supply compared to the vein, but the nerve plexus in the vein displayed a more uniform arrangement. In contrast, relatively few nerve fibres displayed calcitonin gene-related peptide, substance P, or vasoactive intestinal peptide immunoreactivity in either the artery or vein. The calcitonin gene-related peptide immunoreactive fibres had a similar distribution to that of the substance P containing fibres. Using a sensitive in vitro method the vasomotor responses to perivascular peptides were characterized. In the femoral artery NPY potentiated alpha 1-adrenoceptor mediated contractions, and had very little effect by itself. In contrast, 10(-7) M NPY contracted femoral veins by up to 68% relative to 60 mM potassium induced contraction, and there was no potentiation of alpha-adrenoceptor mediated contractions. Acetylcholine, peptide histidine isoleucine, vasoactive intestinal peptide, substance P and calcitonin gene-related peptide, all relaxed the contracted femoral artery and vein. Regarding the putative parasympathetic neurotransmitters, acetylcholine caused stronger relaxation of veins as compared to arteries whereas for vasoactive intestinal peptide and peptide histidine isoleucine the relaxations were stronger in the arterial preparation. These three agonists were more potent in the femoral vein. Substance P was more potent on the femoral vein, having the same maximum response in both preparations. On the other hand, the response induced by CGRP was some three times greater in the venous than in the arterial preparation. These data reveal that although there appear to be only minor differences in the peptidergic innervation of the rat femoral artery and vein pronounced differences occur in the peptide effector responses. The data support the concept that perivascular peptides play different roles in regulating various parts of the circulation.

Comparison of resting and stimulus-evoked catecholamine release from the femoral and renal vascular beds of the dog.

Plasma levels of dopamine (DA) and noradrenaline (NA) were measured in arterial and in femoral and renal venous blood of chloralose-anaesthetised dogs at rest, and during electrical stimulation of the femoral and renal sympathetic nerve supplies. In the femoral bed, sympathetic nerve stimulation elevated venous efflux of NA, but did not reproducibly elevate DA efflux: when this was increased, it comprised less than 1% of the stimulus-evoked catecholamine efflux. By contrast, renal nerve stimulation liberated both NA and DA from the kidney, and DA comprised about 8% of the total stimulus-evoked efflux. Comparison of efflux from intact and denervated kidneys indicated substantial neurogenic release of both NA and DA at rest, with DA comprising 20% of this efflux. The results extend previous evidence for dopaminergic sympathetic innervation of the dog kidney, and suggest that both dopaminergic and noradrenergic renal nerves are tonically active in anaesthetised animals.

Responses of spinal cord neurons following stimulation of A beta femoral-saphenous venous afferent fibers.

A population of large (A beta) afferents is known to have endings in the wall of the femoral-saphenous vein. These afferents project to the lower lumbar spinal cord. The purpose of the present study was to identify, localize, and characterize spinal neurons that receive inputs from such afferents. Responses of 50 neurons in the L6 spinal cord segment of decerebrate-spinal cats or intact cats anesthetized using alpha-chloralose were recorded following electrical stimulation of these afferents. Observations were also made on the convergence of muscle and cutaneous afferent inputs onto neurons driven by stimulation of afferents terminating in the femoral-saphenous vein. All recording sites were marked either by intracellularly staining the element characterized with HRP or by extracellularly iontophoresing a small quantity of this tracer. The cells were driven for long durations (mean of 51.5 ms, S.E.M. of 10.0) by single-shock stimulation of femoral-saphenous venous afferents. The recording sites were located in Rexed's laminae IV-VIII and X. Eight of the 50 neurons were activated by venous afferent stimulation at latencies equal to or shorter than that of the first negative wave of the cord dorsum potential; these units were driven at a mean latency of 1.4 ms (S.E.M. of 0.25) following the arrival of the afferent volley at the cord and were assumed to receive monosynaptic, or at least relatively direct, inputs from the primary afferents. Most of these cells (6 of 8) were located in lamina V. The majority of the neurons studied (37 of 50) were activated at latencies longer than 3 ms following the arrival of the afferent volley at the cord; about half (19 of 37) of those activated at longer latencies were located in lamina VII, and the rest were scattered among the other laminae. Twenty-eight of 40 venous afferent-driven cells tested could also be activated by electrical stimulation of either the posterior tibial or sural nerve. In general, the stimulation intensities necessary to activate the neurons were only sufficient to excite large (A alpha or A beta) muscle and cutaneous afferents. Neurons receiving the shortest latency inputs from the femoral-saphenous vein were less likely to receive convergent inputs from muscle or skin than were neurons activated by venous afferents at longer latency.

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.

Tracing of afferent pathways from the femoral-saphenous vein to the dorsal root ganglia using transport of horseradish peroxidase.

The retrograde transport of horseradish peroxidase (HRP) was used to trace afferents from the femoral-saphenous vein to the dorsal root ganglia in the cat. Afferents arising along the entire length of the vein projected to very localized spinal levels; 63% of the labeled cells counted were located in the L6 dorsal root ganglion, 37% were located in the L5 ganglion and less than 1% were located at other levels. Most of the cell bodies labeled by the application of HRP to the femoral-saphenous vein were small in size (diameter less than 35 microns). However, some large cell bodies (diameter greater than 50 microns) were also noted. It was estimated that over two-thirds of the femoral-saphenous venous afferents were C fibers; at least 15% were estimated to be A fibers. The largest venous afferents were predicted to conduct action potentials at approximately 60 m/s.

Mechanosensitive afferents of femoral-saphenous vein.

Reflex contraction of cat hindlimb skeletal muscles can be induced by mechanical stimulation of afferents in the femoral-saphenous vein. Afferent fibers innervating this vein have been previously reported. Hence, it is evident that mechanoreceptors must be present in the femoral-saphenous vein, yet there have never been direct recordings of sensory nerves that respond to mechanical stimulation of the vein. This study recorded the activity of these venous mechanoreceptors and tested their response to increased intravenous pressure. The activity of single venous afferents was recorded from thin filaments of the saphenous nerve. The afferent response was recorded during external mechanical probing of the vein wall and increases in intravenous pressure. Their venous pressure threshold and adaptation to static pressures were determined. All the afferents responded to both types of mechanical stimuli. Static increases in venous pressure resulted in two types of afferent discharge behavior: slowly adapting and rapidly adapting. Venous pressure thresholds varied considerably with a range of 22.2-174.7 mmHg. The estimated conduction velocities are 2.8-7.8 m/s, suggesting type III, A-delta-myelinated mechanoreceptors. These results demonstrate the presence and mechanical sensitivity of venous afferents that transduce intravenous pressure. Their role in the reflex responses to vein distension is unclear and awaits further investigation.

Effects of temperature on alpha adrenoceptors in limb veins: role of receptor reserve.

In cutaneous veins of the dog, cooling augments the response to sympathetic nerve stimulation and exogenous norepinephrine (NE). The postjunctional alpha adrenoceptors in this blood vessel belong to both the alpha 1 and alpha 2 subtypes. Cooling augments alpha 2-adrenergic responses (presumably because of an increased receptor affinity), but depresses alpha 1-adrenergic responses (presumably because of a direct inhibitory effect on the contractile process). When agonists of high efficacy such as NE or phenylephrine are used, an alpha 1-adrenoceptor reserve is present that buffers the response from the inhibitory effect of cooling. This allows the potentiating effect of cold on the alpha 2-adrenergic component of the response to catecholamines to predominate, and the contractile response to exogenous NE and sympathetic nerve stimulation is augmented. By contrast, in deep veins of the limb, cold reduces the contractions evoked by alpha 1- and alpha 2-adrenergic activation. This can be explained best by the absence of a receptor reserve for alpha 1-adrenergic agonists of high efficacy, combined with a reduced density of postjunctional alpha 2 adrenoceptors.

Properties of femoral venous afferent inputs and lumbosacral distribution of spinal evoked activity.

The present studies were done to determine details of the anatomical and physiological characteristics of femoral-saphenous venous afferent input to the lumbar spinal cord. Gross anatomical examination revealed that afferent bundles could be seen coursing from the saphenous nerve to the femoral-saphenous vein. Compound action potentials elicited by femoral-saphenous venous afferent stimulation were recorded from the femoral nerve and in dorsal rootlets of the 6th lumbar cord segment. The compound action potentials included activity correlated with that of fibers conducting impulses at the rate of 31 to 61 m/s. Lumbar cord dorsum potentials elicited by femoral-saphenous venous afferent stimulation were abolished by rhizotomy of the most caudal rootlets of the 6th lumbar cord segment. L6 was also the cord segment from which the largest amplitude cord dorsum negative potentials were recorded, while action potentials with large late positive waves were recorded from more caudal cord segments. These observations suggested that the L6 segment contained the largest number of spinal neurons responding to primary femoral-saphenous venous afferent input, and that input reached the more caudal segments via intersegmental connections. A proposed physiological role of these afferents is briefly described.

Activation of spinal cord interneurons which process inputs from the femoral-saphenous vein.

Recordings were made from single spinal cord interneurons which could be activated by electrical stimulation of afferents terminating in the wall of the femoral-saphenous vein. Interneurons were either excited or both excited and inhibited by venous afferent stimulation. Most of the venous afferent-driven interneurons could also be driven by electrical activation of A-alpha beta muscle and cutaneous afferents. Stimulation of several different muscle nerves drove single interneurons.

Properties of spinal cord processing of femoral venous afferent input revealed by analysis of evoked potentials.

Two characteristics of spinal cord interneurons which receive inputs from femoral-saphenous venous afferents were examined. The shortest pathway between primary femoral-saphenous venous afferents and alpha-motoneurons was shown to be a di- or tri-synaptic circuit. In addition, the largest focal synaptic field potentials elicited by venous afferent stimulation at short latency were recorded from Rexed's lamina V. It was thus concluded that most of the first interneurons excited by venous afferents are found in this lamina.

Dopamine preferentially stimulates postsynaptic alpha 2-adrenoceptors in the femoral vascular bed, but alpha 1-adrenoceptors in the renal vascular bed of the anaesthetised dog.

The decrease in blood flow in response to dopamine (DA) injected intraarterially (i.a.) into the femoral or renal vascular beds was examined in the anaesthetised dog. DA or noradrenaline (NA) were 10 times more potent as vasoconstrictor agents in the femoral than in the renal vasculature. In the femoral bed, the DA induced vasoconstriction was completely resistant to antagonism by prazosin (30-300 micrograms/kg i.v.), but was dose-dependently blocked by the alpha 2-receptor antagonist idazoxan (30-300 micrograms/kg i.v.). In the renal bed the vasoconstrictor effects of DA were resistant to blockade by idazoxan, but were prazosin sensitive indicating that alpha 1-adrenoceptors were involved in this response. The alpha-receptor agonist profile for DA was not modified in the femoral bed after blockade of dilatory D1-receptors with SCH 23390 (0.5 mg/kg i.v. and 0.1 mg/kg per h i.v.). However, this antagonist significantly increased the vasoconstrictor potency for DA in the renal bed. The decrease in femoral blood flow induced by an injection of DA, appears to be mediated by alpha 2-adrenoceptors. In the renal vascular bed where the predominant alpha-adrenoceptor corresponds to the alpha 1-subtype and there are few postsynaptic alpha 2-receptors subserving vasoconstriction, DA can stimulate alpha 1-receptors but this action requires higher doses of agonist than those needed for alpha 2-adrenoceptor stimulation.

Vascular adrenergic neuroeffector function does not decline in aged rats.

To investigate adrenergic control of blood vessels during aging, rats aged 6, 12, 20, and 27 months were studied using in vitro techniques. Accumulation of [3H]norepinephrine, one index of adrenergic nerve density, did not alter with age in the femoral or renal arteries or renal vein. In the femoral vein [3H]norepinephrine accumulation was greater at 6 and 27 months of age. Norepinephrine sensitivity was determined in both an innervated vessel, the femoral artery, and a non-innervated vessel, the carotid artery. In both cases, sensitivity to norepinephrine did not alter with age. In the renal and femoral arteries and veins, no significant changes in maximum responses to norepinephrine (10(-5) M), potassium chloride, or transmural nerve stimulation were seen with advancing age. Furthermore, frequency response curves (2-16 Hz, 200 pulses) did not differ with age for any of the four vessels studied, with one exception. The response to stimulation at 4 Hz of the femoral vein from 6-month-old rats was significantly larger than responses at other ages. During nerve stimulation, the renal vein exhibited rapid contractions superimposed upon the maintained contractile response. This type of rapid contraction occurred only rarely (1 out of 5) in the renal vein from 27-month-old rats. In summary, neither adrenergic nerve density as reflected by [3H]norepinephrine accumulation nor norepinephrine sensitivity decline with age. As the net effect of various components, the ability of vascular smooth muscle to respond to adrenergic nerve stimulation is also maintained during advancing age.

Interactions between femoral venous afferents and lumbar spinal reflex pathways.

This study reports findings of spinal reflex connections of afferent fibers electrically excited in the wall of the femoral vein. Condition-test experiments, and EMG recordings revealed that the femoral venous afferents have facilitatory connections to flexor and extensor motoneurons of both the proximal and the distal hindlimb muscles. Femoral venous afferent stimulation which produced facilitation, also produced inhibition of the test reflexes following the facilitation. Because the inhibition was enhanced by diazepam injection and because the inhibitory time-course correlated closely to the time-courses of both dorsal root potentials and individual tests of primary afferent depolarization, the inhibition was suggested to be produced by presynaptic inhibition. The potentially significant role of the venous afferent connections in a reflex-elicited skeletal muscle pump or in an increase in intramuscular venous counterpressure is discussed.