The efficiency of mesenchymal cell intracerebral transplantation for corrections of cerebral microcirculation age-related alterations in rats

Using television-based vital microscopy and immunohistochemical analysis, we have assessed the effect of syngeneic mesenchymal stem-cell (MSC) intracerebral transplantation on the brain cortex structure and microcirculation in the pia mater of old rats. Setting up the “open field” system, we studied the effect of MSC transplantation on the position-finding and discovery behavior of old animals. We found that the density of microvascular network of the pia mater increased by about 1.9-fold in MSC recipients compared to age-matched controls. The density of the arteriolar area of microvascular network of the pia mater approximately doubled. The reactivity of newly developed arterioles was nearly equal to that of native microvessels. The intracerebral transplantation procedure was itself traumatic for rat brain cortex, but it did not affect microcirculation in the contralateral hemisphere. Intracerebral transplantation of MSCs did not improve the locomotor behavior and emotional stage of old rats; neither did it increase their position-finding and discovery activity.     

The effect of intracerebral mesenchymal stem cells transplantation on the density of microvascular network of the pial matter of the rat brain cortex

Using a TV installation for studying the microcirculation (with 30-160-fold magnification), the density of microvascular network in the pia matter of the rat brain sensomotor cortex was determined after intracerebral transplantation of mesenchymal stem cells (MSC) or (as control) of the MSC cultivation nutrition medium, or of saline. The results have shown that intracerebral transplantation does not change density of microvascular network in the pia mater of the ipsilateral hemisphere. Transplantation of the MSC led to a 1.8-fold increase of density of the pia matter of the contralateral hemisphere as compared with control animals; the number of arterioles in the same zone was 2.5-fold higher than in intact rats.     

Control of oxidative stress in microcirculation of spontaneously hypertensive rats

One mechanism for organ damage in individuals with arterial hypertension may be due to oxygen free radical production. This study was designed to localize free radicals in a microvascular network of mature spontaneously hypertensive rats (SHRs) and normotensive Wistar-Kyoto (WKY) rats. Because glucocorticoids play a role in pressure elevation of SHRs, we investigated their role in microvascular free radical formation. Oxygen radical production in mesentery was detected by tetranitroblue tetrazolium reduction to formazan aided by digital light-absorption measurements. Formazan deposits were observed in the endothelial cells and lumens of all microvessels and in lymphatic endothelia but were fewer in tissue parenchyma. The formazan distribution in younger (14-16 wk old) WKY rats and SHRs was heterogeneous with low values in capillaries and small arterioles/venules (<30 microm) but enhanced deposits in larger venules. Adrenalectomy served to reduce the formazan density in SHRs to the level of WKY rats, whereas dexamethasone supplementation of the adrenalectomized rats caused elevation in the larger venules of SHRs. In older (40 wk old) SHRs, formazan levels were elevated in all hierarchies of microvessels. After pressure reduction was employed with chronic hydralazine treatment, the formazan deposits were reduced in all locations of the microcirculation in both WKY rats and SHRs. Elevated formazan deposits were also found in lymphatic endothelium. These results suggest that oxygen free radical production is elevated in both high- and low-pressure regions of SHR microcirculation via a process that is controlled by glucocorticoids. Older SHRs have higher formazan levels than younger SHRs in all microvessels. Chronic hydralazine treatment, which serves to reduce arterial blood pressure, attenuates tetranitroblue tetrazolium reduction in WKY rats and SHRs even in venules of the microcirculation, which has no micropressure elevation. Free radical production may be a more global condition in SHRs and may not be limited to arteries and arterioles.     

Prolonged tissue PO2 reduction after contraction in spinotrapezius muscle of spontaneously hypertensive rats

We tested the hypothesis that a deficit in oxygen extraction or an increase in oxygen demand after skeletal muscle contraction leads to delayed recovery of tissue oxygen tension (Po(2)) in the skeletal muscle of hypertensive rats compared with normotensive rats. Blood flow and Po(2) recovery at various sites in the spinotrapezius muscle of spontaneously hypertensive rats (SHRs) were evaluated after a 3-min period of muscle contraction and were compared with corresponding values in Wistar-Kyoto rats (WKYs). The recovery of tissue Po(2) [75 +/- 7 (SHRs) vs. 99 +/- 12% (WKYs) of resting values] and venular Po(2) [72 +/- 13 (SHRs) vs. 104 +/- 10% (WKYs) of resting values] were significantly depressed in the SHRs 30 s postcontraction. The delayed recovery persisted for 120 s postcontraction for both tissue [86 +/- 11 (SHRs) vs. 119 +/- 13% (WKYs) of resting values] and venular [74 +/- 2 (SHRs) vs. 100 +/- 9% (WKYs) of resting values] Po(2) levels. There was no significant difference in the recovery of arteriolar Po(2) between the two groups 30 s postcontraction [95 +/- 7 (SHRs) vs. 84 +/- 8% (WKYs) of resting values]. Values for resting diameter of arcade arterioles in the two groups were not different [52 +/- 3 (SHRs) vs. 51 +/- 3 microm (WKYs)], but the arteriolar diameter after the 3-min contraction period was greater in the SHRs (71 +/- 4 microm) than the WKYs (66 +/- 4). Likewise, red blood cell (RBC) velocity [5.8 +/- 0.3 (SHRs) vs. 4.7 +/- 0.2 mm/s (WKYs)] and blood flow [23.0 +/- 0.8 (SHRs) vs. 16.0 +/- 1.0 nl/s (WKYs)] measurements were significantly greater in the SHRs at 30 s postcontraction. The delayed recovery of tissue Po(2) in the SHRs compared with the WKYs can be explained by a decrease in oxygen diffusion from the rarefied microvascular network due to the increased RBC velocity and the shorter residence time in the microcirculation and the consequent disequilibrium for oxygen between plasma and RBCs. The delayed recovery of venular Po(2) in the SHRs is consistent with this explanation, as venular Po(2) is slowly restored to baseline by release of oxygen from the RBCs. This leaves the arterioles in the primary role as oxygen suppliers to restore Po(2) in the tissue after muscle contraction.     

Postnatal physiological development of rats after acute prenatal hypoxia

The aim of study was to investigate the physiological development of the brain and behaviour in rats subjected to prenatal hypoxia on the 13.5th day of embryogenesis. We have found that such rats manifested a delayed physiological development and a change in nervous tissue of the sensorimotor cortex, as well a disturbed formation of motor responses during the first month of postnatal ontogenesis. During maturation these modifications were in part compensated, however we observed a decrease of the rats' ability to learn new forepaw movements. The destruction of the brain tissue and the modification of neurons composition in the sensorimotor cortex correlated with changes of behaviour at different stages of ontogenesis. Thus, changes of the conditions under which an organism develops during embryogenesis, predetermine a disturbance in ontogenesis and the learning ability.     

Results of transplanting regions of the midbrain of human embryos into the brain of adult rats

A principle possibility for transplantation of the midbrain areas of the human embryos into the brain of mature rats has been demonstrated. For successful xeno-transplantations the midbrain of 7-8-week-old embryos is the most suitable. The intracerebral transplants grow predominantly along the capsule of the white substance fasciculi. Cell differentiation of the intraventricular transplants in comparison with the intramedullary ones takes less time. Manifestation of the glial scar at the borders of the transplant depends on its localization. Perspectives on applying the method of the nervous embryonal tissue transplantation into the brain of mature animals for reconstruction and formation of new centers in order to compensate cerebral defects are discussed.     

Thalidomide decreases intrahepatic resistance in cirrhotic rats

Increased intrahepatic resistance (IHR) within cirrhotic liver is caused by increased endotoxemia, cytokines tumor necrosis factor-α (TNF-α), vasoconstrictor thromboxane A₂ (TXA₂), and disrupted microvasculatures. We evaluated the effects of thalidomide-related inhibition of TNF-α upon the hepatic microcirculation of cirrhosis in rats. Portal venous pressure (PVP), hepatic TNF-α, expression of thromboxane synthase (TXS), and leukocyte common antigen (LCA) were measured in bile-duct-ligated (BDL) rats receiving 1 month of thalidomide (BDL-thalido rats). Portal perfusion pressure (PPP), IHR, and hepatic TXA₂ production were measured in the isolated liver perfusion system. Intravital microscopy was used to examine hepatic microvascular disruptions. In BDL-thalido rats, PVP, PPP, IHR, hepatic TXA₂ and TNF-α, hydroxyproline content, expression of TXS and LCA, and LPS-induced leukocyte recruitment were significantly decreased. Conversely, hepatic microvascular density and perfused sinusoids were significantly increased. Thalidomide decreased PVP and IHR by reducing hepatic TXA₂ and improving hepatic microvascular disruptions in rats with biliary cirrhosis.     

Homotopic transplantation of embryonic neocortical tissue into the brain of adult rats after its damage

Structural characteristics (survival, growth, connections) have been studied in the transplant of the cerebral cortex tissue in Wistar rat embryos (18-day-old), implanted into the brain of mature rats of the same line at various time after a partial lesion of the sensomotor cortex. In 3-5 months after transplantation the light microscopy methods demonstrate that spatial interconnections of the transplant and the injured brain of the recipient depend on time interval between the cerebral lesion and transplantation of the embryonal nervous tissue. Horseradish peroxidase (HP) is ionophoretically injected into the recipient's cerebral tissue away from the place of transplantation. In the transplant retrogradely labelled HP neurons are revealed. This demonstrates efferent connections of the implanted tissue with the host's brain. Presence of the anterogradely labelled nervous terminals in the transplant tissue demonstrates existence of afferent connections of the transplant with the recipient's tissue. Possible mechanisms of survival, growth and formation of connections of the transplant in the injured brain of the mature animal are discussed.     

Mathematical model of the dynamics of transport of inert gases in the microcirculatory system

A mathematical model imitating transport of inert gases in the system of microcirculation under increased pressures was constructed. It has been shown that saturation of microareas nucleus of the brain cortex of average dimensions proceeds in about 90 sec. Effect of the blood flow velocity, gases tension in arterial blood and density of the capillary net on the dynamics of mass transfer of gases in a tissue was investigated.     

Low-flow vascular remodeling in the metabolic syndrome X

Peripheral microvascular dysfunction is a common affliction in patients with the metabolic syndrome X. Previous studies have described a number of vascular impairments in vasomotor control in both human patients and animal models of syndrome X, but the net effect of these impairments on microvascular structure has not been examined. The goal of the current study was to test the hypothesis that syndrome X reduces muscle perfusion and induces vascular remodeling. The obese Zucker rat was used as a model of syndrome X, and the microcirculation of the hindlimb and brain were examined. Obese Zucker rats were obese, hyperlipidemic, hyperinsulinemic, and hyperglycemic. Blood flow to the hindlimb was reduced by 59% in obese rats relative to lean rats. Skeletal muscle resistance arteries of the hindlimb microcirculation of obese rats had thinner walls, smaller lumens, and reduced distensibility. Hindlimb microvessels from obese rats also demonstrated reduced expression of vascular smooth muscle cell markers. Each of these traits is consistent with low-flow remodeling. In contrast, the cerebral microcirculation, where flow is vigorously autoregulated, showed no vascular remodeling nor were there changes in microvascular smooth muscle marker expression. Neither physical activity nor muscle mass were significantly different between lean and obese rats. Taken together, these findings suggest that syndrome X, by reducing hindlimb blood flow, induces a marked remodeling of microcirculation to favor smaller, less distensible vessels. This remodeling may result in an architectural limitation of maximum perfusion capacity and may be an important maladaption in the progression of peripheral microvascular disease.     

Microvascular oxygen distribution in awake hamster window chamber model during hyperoxia

The microvascular effects and hemodynamic events following exposure to normobaric hyperoxia (because of inspiration of 100% O2) were studied in the awake hamster window chamber model and compared with normoxia. Hyperoxia increased arterial blood Po2 to 477.9 +/- 19.9 from 60.0 +/- 1.2 mmHg (P < 0.05). Heart rate and blood pressure were unaltered, whereas cardiac index was reduced from 196 +/- 13 to 144 +/- 31 ml.min-1.kg-1 (P < 0.05) in hyperoxia. Direct measurements in the microcirculation showed there was arteriolar vasoconstriction, reduction of microvascular flow (83% of control, P < 0.05), and functional capillary density (FCD, 74 +/- 16% of control), the latter change being significant (P < 0.05). Calculations of oxygen delivery and oxygen consumption based on the measured changes in microvascular blood flow velocity and diameter and estimates of oxygen saturation corrected for the Bohr effect due to the lowered pH and increased Pco2 showed that oxygen transport in the microvascular network did not change between normal and hyperoxic condition. The congruence of systemic and microvascular hemodynamics events found with hyperoxia suggests that the microvascular findings are common to most tissues in the organism, and that hyperoxia, due to vasoconstriction and the decrease of FCD, causes a maldistribution of perfusion in the microcirculation.     

Afferent and efferent connections of cortical transplants implanted into the damaged sensorimotor area of the cerebral cortex of mature rats

Afferent and efferent connections of the transplant, implanted in the previously damaged sensorimotor area of the mature rat cerebral cortex have been studied by means of axonal transport of horseradish peroxidase. For 5 months after transplantation neural axons of the transplant are capable to reach the caudo-putamen and thalamic structures, while connections with the spinal cord are absent. The afferent connections of the transplant are minimal and belong only to the neighbouring areas of the cortex and the caudo-putamen of the recipient brain. Presence of efferent projections to the striate and thalamic structures demonstrates specificity of the projections formed; this can be a morphological base for restoration of the functions lost after the damage of the sensorimotor area of the cortex in mature animals.     

Portal branch ligation induces a hepatic arterial buffer response, microvascular remodeling, normoxygenation, and cell proliferation in portal blood-deprived liver tissue

Portal branch ligation (PBL) may prevent liver failure after extended hepatic resection. However, clinical studies indicate that tumors within the ligated lobe develop accelerated growth. Although it is well known that tumor growth depends on the host's microvascularization, there is no information about how PBL affects the hepatic microcirculation. Our aims were to determine hepatic artery response, liver microcirculation, tissue oxygenation, and cell proliferation after PBL. Therefore, we used intravital multifluorescence microscopy, laser-Doppler flowmetry, immunohistochemistry, and biochemical techniques to examine microcirculatory responses, microvascular remodeling, and cellular consequences after left lateral PBL in BALB/c mice. During the first 7 days, PBL induced a reduction of left hilar blood flow by approximately 50%. This resulted in 80% sinusoidal perfusion failure, significant parenchymal hypoxia, and liver atrophy. After 14 days, however, left hilar blood flow was found to be restored. However, remodeling of the microvasculature included a rarefaction of the sinusoidal network, however, without substantial perfusion failure, compensated by a hepatic arterial buffer response and significant sinusoidal dilatation. This resulted in normalization of tissue oxygenation, indicating arterialization of the ligated lobe. Interestingly, late microvascular remodeling was associated with increased endothelial nitric oxide synthase expression, significant hepatocellular proliferation, and weight gain of the ligated lobe. Thus PBL induces only an initial microcirculatory failure with liver atrophy, followed by a hepatic arterial buffer response, microvascular remodeling, normoxygenation, and hepatocellular proliferation. This may explain the accelerated tumor progression occasionally observed in patients after PBL.     

High energy phosphate compounds and ATPase activity of mitochondria in the brain of rats of different ages

Adenosine phosphate and creatine phosphate amount was determined in the brain tissue of 3-4-week, 6-8 month and 26-26 month old mongrel female rats. The maximum ATP and creatine phosphate amount and the minimum of ADP and AMP were found in young rats. In adult rats as compared with the young the ATP amount is the same, the ADP and AMP level rises, that of creative phosphate falls and energy charge decreases. In the brain of old rats the ATP and ADP amount falls, that of creatine phosphate and AMP remains at the level of mature-age animals. Despite a decrease in the ATP amount in the brain at the old age, the Mg, DNP- ATPase activity of mitochondria isolated from brain cortex and stem of the old rats remains at the level typical of adult animals.     

Age-related changes in receptor-mediated phosphoinositide hydrolysis in various regions of rat brain.

The effects of age on cholinergic markers and receptor-stimulated phosphoinositide hydrolysis was examined in the frontal cortex and striatum of male Fischer-344 rats. Choline acetyltransferase activity was decreased 27% in the striatum of aged (24 month) rats compared to young (3 month) controls. Muscarinic receptor density as measured by [3H]-quinuclidinyl benzilate binding showed a similar 26% decrease in the striatum of aged rats. Phosphoinositide hydrolysis was measured by the release of inositol phosphate (IP) from tissue slices prelabeled with [3H]myoinositol in response to carbachol, norepinephrine, and quisqualate. In the cortex, stimulated IP release was significantly greater in slices from aged rats compared to young rats for all three agonists. In contrast, stimulated IP release was significantly decreased in striatal slices from aged rats compared to young for all three agonists. These data indicate a differential effect of age on agonist-stimulated phosphoinositide hydrolysis in the cortex and striatum. The decreased responsiveness in the latter area may result from the age-related loss of postsynaptic receptors.     

Effect of transplantation of embryonic neural tissue on the dynamics of brain edema following experimental craniocerebral trauma

The effect of transplantation of embryonic neural tissue (ENT) on the dynamics of brain edema following heavy craniocerebral trauma (CCT) made to the left parietotemporal area was studied in rats. The brain tissue impedance was measured in the damaged and contralateral hemispheres 1 to 30 days after the trauma in the animals of three groups: (i) without any procedures after the CCT, (ii) with surgical treatment of the damaged brain area, and (iii) with transplantation of 1–2 mm³ sensorimotor cortex fragments from 18-day-old rats grafted into the cavity created by this treatment. At the first day after CCT, the impedance in the damaged hemisphere decreased by 30–37%, while the impedance in the contralateral hemisphere decreased approximately by 20%, compared with the control, which was evidence of the development of intensive generalized edema. In the group of animals with the ENT transplantation, the edema involuted noticeably faster than in the other two groups: the mean impedance value reached 97.9% of the control value (before the damage) already by the 7th post-traumatic day. Complete recovery of the impedance by the 30th day was observed only in the animals with transplantation. The adequacy of an impedancemetric technique for investigation of the dynamics of water-electrolyte re-distribution in the brain tissue, and the mechanisms underlying corrective effect of ENT transplantation on the edema dynamics in the post-traumatic period are discussed.     

Electron microscopic and cytointerferometric study of the cortical neurons from the 2d-generation progeny of preneurosensitized female rats

A study was made of the sensorimotor cortex of the brain of the second generation offspring of preneurosensitized female rats. Vacuolization of the neurocyte nuclei, elevated lability of nuclear membranes, appearance of numerous dark neurons were discovered at all times of postnatal ontogenesis (from 2 to 90 days). At the same time ultrastructure of a considerable number of neurons was unchanged. The tendency toward normalization of cellular structures was noted by the 3d month. The one-month-old rats demonstrated a decrease in the content of protein substances in the nucleus and cytoplasm of the neurons as compared to normal. By the 3d month this indicator rose but did not reach the control level. It is concluded that neurosensitization of females before pregnancy affects the morphofunctional state of the neurons of the cerebral cortex in both first and second generation offspring, although the changes seen in the latter offspring are less marked, being compensated for with time.     

Effect of intracerebral implantation of the embryonicLocus Coeruleus on the conditioned reflex avoidance reaction in rats with experimental atrophy of the frontotemporal brain cortex

We studied the effect of intracerebral grafting of the embryoniclocus coeruleus from donor rats on the conditioned reflex emotional avoidance reaction (EAR) and contents of adrenaline and noradrenaline in the brain structures and body tissues of recipient rats. Group 1 of the animals consisted of intact rats, group 2 included rats with electrolytic impairment and subsequent atrophy of the frontotemporal brain cortex regions, while in group 3 the rats with impaired cortex were subjected to parallel transplantation of the embryoniclocus coeruleus into the injured region. Group-3 animals manifested a considerable level of recovery of the conditioned reflex behavioral stereotype disturbed after the cortical injury. Biochemical analysis showed that transplantation of the embryoniclocus coeruleus exerts a rehabilitative/regulating influence on the functioning of the catecholaminergic systems in the group-3 rats.     

Distribution of microspheres in the brain of hypertensive rats

The blood perfusion of different parts of the brain tissue was examined by means of microspheres 15 and 50 micron in diameter, in normotensive control rats and in animals with experimental renovascular hypertension. The microspheres were labelled with fluorescein isothiocyanate and their numbers in the tissue were determined in consecutive histological sections by UV microscopy. In the control rats, the incidence of wedged microspheres per 1 mm3 tissue was high in the cerebellum, cerebral cortex, subcortical tissue and pons Varolii, but low in the thalamic and hypothalamic regions, indicating that these parts were relatively poorly perfused with blood. The significantly greater accumulation of microspheres in the cortex and subcortical tissue of hypertensive rats seems to have been due to hypertensive narrowing of the arterioles. Conversely, the diminished incidence of microspheres in the thalamus and hypothalamus may have been due partly to microsphere trapping in the narrowed upstream blood vessels and partly to thinning of the capillary network. Total microsphere recovery in the brains of the control and the hypertensive rats was almost identical, implying that only the distribution of brain blood perfusion is altered in experimental hypertension.     

Sensorimotor experience influences recovery of forelimb abilities but not tissue loss after focal cortical compression in adult rats

Sensorimotor activity has been shown to play a key role in functional outcome after extensive brain damage. This study was aimed at assessing the influence of sensorimotor experience through subject-environment interactions on the time course of both lesion and gliosis volumes as well as on the recovery of forelimb sensorimotor abilities following focal cortical injury. The lesion consisted of a cortical compression targeting the forepaw representational area within the primary somatosensory cortex of adult rats. After the cortical lesion, rats were randomly subjected to various postlesion conditions: unilateral C5-C6 dorsal root transection depriving the contralateral cortex from forepaw somatosensory inputs, standard housing or an enriched environment promoting sensorimotor experience and social interactions. Behavioral tests were used to assess forelimb placement during locomotion, forelimb-use asymmetry, and forepaw tactile sensitivity. For each group, the time course of tissue loss was described and the gliosis volume over the first postoperative month was evaluated using an unbiased stereological method. Consistent with previous studies, recovery of behavioral abilities was found to depend on post-injury experience. Indeed, increased sensorimotor activity initiated early in an enriched environment induced a rapid and more complete behavioral recovery compared with standard housing. In contrast, severe deprivation of peripheral sensory inputs led to a delayed and only partial sensorimotor recovery. The dorsal rhizotomy was found to increase the perilesional gliosis in comparison to standard or enriched environments. These findings provide further evidence that early sensory experience has a beneficial influence on the onset and time course of functional recovery after focal brain injury.