Vascular response to radiation injury in the rat lung.

Changes in relative left-to-right lung blood flow ratios were followed as an index of vascular radiation injury in left-hemithorax-irradiated Sprague-Dawley rats. Single doses of 11 to 21 Gy gamma radiation resulted in a dose-dependent decrease in relative blood flow to the irradiated lung from 3 to 5 weeks after exposure during the development of pneumonitis. Blood flow returned to near normal by 5 weeks after lower doses (11-13.5 Gy). After a single dose of 15 Gy the left-to-right blood flow ratio recovered to 75% of normal at 12 weeks and leveled off. Following 18 Gy irradiation a second period of reduced flow began 16 weeks after exposure. After 21 Gy irradiation flow to the irradiated side remained low for 1 year after exposure. Rats that received a single dose of 18 Gy to the left hemithorax were also treated with one or two of the following drugs: captopril, cyproheptadine, dexamethasone, diethylcarbamazine, penicillamine, or theophylline. Dexamethasone was most effective at preventing the decrease in blood flow to the irradiated lung when treatment was continued through the pneumonitis period and dose was not tapered until 8 weeks after radiation exposure. All other drugs and drug combinations were, for the most part, virtually ineffective after the pneumonitis period. There was a relatively poor correlation with earlier vascular permeability surface area product studies. This suggests that endothelial damage, as well as damage to other cell types, contributes to the development of post-irradiation fibrosis in the lung.     

Epithelial response of the rat gastric mucosa to chronic superficial injury.

Chronic injury to the healthy gastric mucosa with noxious agents such as aspirin or alcohol induces a progressive strengthening of the stomach wall against these insults. The present study examined the histologic response of the rat gastric mucosa to chronic destruction of the superficial mucosa for one month with hypertonic saline. The number, position and morphology of proliferating, parietal, G and D cells were followed during mucosal injury and one month of recovery. The results showed that chronic injury reduced parietal cell numbers by about 30 percent, particularly in the middle of the mucosal thickness where a clear zone was formed by hypertrophy of mucous neck-like cells. G cells were also reduced by about 50 percent, but there were no changes in D cells. Chronic injury induced a marked increase in the number of antral (+112 percent) and fundic (+250 percent) proliferating cells. CONCLUSION: The rat gastric mucosa responds to chronic superficial injury by down-regulation of acid secretory cells and gastrin secreting cells and an up-regulation of proliferating cells. The appearance of a prominent layer of mucous neck-like cells may indicate a new secretory function for these cells.     

Lateralization of catecholaminergic and behavioral response to cerebral infarction in the rat.

The effects of left middle cerebral artery ligation on brain catecholamine concentrations and behavior were compared with those effects of right middle cerebral artery ligation which have previously been reported. Although the lesion caused by middle cerebral artery ligation appeared to be identical on the two sides, the animals with left hemispheric lesion showed no postoperative change in either mean spontaneous 24 hour activity or mean catecholamine concentrations in several areas of the brain. In marked contrast, as we have previously reported and was confirmed in the present study, the animals with a right hemispheric infarct are hyperactive for about 2 to 3 weeks after surgery and there is a significant decrease in mean norepinephrine concentrations in several brain regions. These studies have demonstrated a remarkable asymmetry in the behavioral and biochemical response to cerebral cortical infarction. It is uncertain whether this asymmetry reflects an underlying hemispheric difference in catecholaminergic or non-catecholaminergic neurons.     

Uptake of Ga-67 into rat cerebral hemisphere and cerebellum. Comparison with Fe-59.

Transferrin and transferrin receptors play an important role in the transport of iron into the brain. To determine whether gallium enters the brain by the same mechanism, uptakes of Ga and 59Fe have been compared under controlled conditions. Rates of gallium penetration into brain (K) were four times slower than those for 59Fe. Kin for Ga when infused with citrate were 0.88 ± 0.24 and 0.94 ± 0.39 x 10 ml gh for cerebral hemisphere and cerebellum, respectively. When infused as the transferrin complex, Ga uptake into the brain was not different from that when infused with citrate. The presence of the anti-transferrin receptor antibody OX-26 significantly reduced uptake of Fe by 60% and 64% into cerebral hemisphere and cerebellum, respectively. By contrast, pretreatment of rats with OX-26 enhanced the uptake of Ga into brain, particularly when infused with citrate; mean increases in uptake of Ga were 120% and 144% for cerebral hemisphere and cerebellum, respectively. Purified Ga-transferrin was also taken up into both brain regions examined in the presence of OX-26. These results indicate that the transport of non-transferrin bound gallium is an important mechanism for gallium uptake into brain.     

The threshold response of neonatal rat cardiomyocytes to gradual enhancement of cryptosporidial invasion

One common cause of excessive cardiac loading in children is infectious gastroenteritis that produces malabsorption and tachyarrythmia. Our recent studies have shown that neonatal cryptosporidial gastroenteritis causes the long-term pathology of cardiomyocytes. In the present work, we studied how cryptosporidiosis of different degrees of severity is reflected on the heart anatomy and polyploidization and remodeling of cardiomyocytes of neonatal rats, as well as on the expression of their gene encoding factor sensitive to hypoxia (HIF-1α). By methods of Real-Time PCR, cytometry, immunocytochemistry, image analysis, and visual study of interatrial septum, we have established that gradual enhancement of infestation by cryptosporidiosis is accompanied by threshold changes in heart. With weak infection, the interatrial septum preserves integrity, while changes in cardiomyocytes are absent. With moderate and strong infection, the changes are expressed approximately equally: in the interatrial septum, foramen ovale appears and the heart becomes atrophied and elongated, while cardiomyocytes lose protein and become thinned and hyperpolyploid. In addition, an excessive level of mRNA of gene HIF-1α appears in the myocardium. It is important to note that the threshold response to the gradual increase of the action is a criterion of triggering of the developmental programming of cardiovascular diseases, as such response is based on irreversible disturbance of the anatomy of the organ and failure of cell differentiation. In our case, the disturbance of the heart is the patent foramen ovale in the interatrial septum, while the disturbance of differentiation is hyperpolyploidization of cardiomyocytes. Our data may be of interest to physicians, as they show for the first time a connection between gastroenteritis, disturbance of the integrity of the interatrial septum, and pathological alterations in cardiomyocytes.     

Sympathetically mediated hypermetabolic response to cerebral ischemia in the rat

Cerebral ischemia, induced in rats by occlusion of the middle cerebral artery resulted in infarcts affecting the basal ganglia and adjacent frontoparietal cortex. Resting oxygen consumption was similar for sham-operated and ischaemic rats immediately after surgery but was elevated in the latter group (peak value 18-21% above controls) 5-6 h post occlusion. By 24 h, these values had returned to control levels. The increase in VO2 was inhibited by injection of the beta-adrenergic antagonist propranolol but was unaffected by injection of the cyclooxygenase inhibitor ibuprofen. The thermogenic activity of brown adipose tissue was assessed from in vitro binding of guanosine diphosphate to mitochondria isolated from intact and surgically denervated lobes of sham-operated and ischemic rats, 6 h after surgery. Brown adipose tissue specific guanosine diphosphate (GDP) binding was elevated by 86% in intact tissue from ischemic compared with sham-operated rats but was identical in denervated tissue from the two groups. Brown adipose tissue activity correlated with resting oxygen consumption in the ischemic group (r = 0.85, p less than 0.01) but not in controls (r = -0.35, NS). Thus occlusion of the middle cerebral artery in the rat may provide a representative model for both stroke and head injury in man. It is associated with a transient increase in metabolic rate and by sympathetically mediated activation of brown adipose tissue in the rat.     

Angiotensin II and the fibroproliferative response to acute lung injury

Angiotensin II (ANG II), generated by activation of local renin-angiotensin systems, is believed to play an important role in tissue repair and remodeling, in part via transforming growth factor-beta (TGF-beta). Angiotensin-converting enzyme (ACE) inhibitors have been shown to abrogate experimental lung injury via a number of potential mechanisms; however, the potentially fibroproliferative role for ANG II in the lung has not been characterized. We hypothesized that, after lung injury, ANG II would stimulate fibroblast procollagen synthesis and promote lung collagen deposition in rats. In vitro, ANG II was a potent inducer of procollagen production in human lung fibroblasts via activation of the type 1 receptor and, at least in part, via the autocrine action of TGF-beta. After bleomycin-induced lung injury, an increase in lung ANG II concentration was observed by day 3 that preceded increases in lung collagen and was maintained until death at day 21. Administration of an ACE inhibitor (ramipril) reduced ACE activity, ANG II concentration, TGF-beta expression, and collagen deposition. Losartan (an ANG II type 1 receptor antagonist) also attenuated the increase in TGF-beta expression and lung collagen deposition. These observations suggest that ANG II, possibly generated locally within the lung, may play an important role in the fibrotic response to acute lung injury, at least in part via the action of TGF-beta. ACE inhibitors and receptor antagonists, already widely used clinically, should be assessed as potential new therapies for fibrotic lung disease.     

Lesion-induced proliferation of vimentin-positive astrocytes in the mouse cerebral hemisphere.

Proliferative response of vimentin expressing astrocytes to injury of the cerebral hemisphere was examined in mouse using 3H-thymidine autoradiography. Within the injury region the vimentin-positive astrocytes proliferation began on the 2nd day after injury, reached its maximal intensity on day 4 and its last evidence was recorded on the 8th postoperative day. The pattern of changes in the distribution of reactive proliferation of the astrocytes was also presented.     

Immunocytochemical studies of astrocytes following injury to the cerebral cortex of the rat

The morphological change of cerebral cortex astrocytes from protoplasmic to glial fibrillary acidic protein (GFAP)-containing cells is induced by injury. Protoplasmic astrocytes that contain no detectable amount of GFAP become filled with GFAP and their processes extend to form the glial scar around the wound. It is hypothesized that this transformation is induced by cAMP and neurotransmitters released from damaged neuronal cells. A similar mechanism may be present in other brain regions following injury or disease.     

Development of the ACTH and corticosterone response to acute hypoxia in the neonatal rat

Acute episodes of severe hypoxia are among the most common stressors in neonates. An understanding of the development of the physiological response to acute hypoxia will help improve clinical interventions. The present study measured ACTH and corticosterone responses to acute, severe hypoxia (8% inspired O(2) for 4 h) in neonatal rats at postnatal days (PD) 2, 5, and 8. Expression of specific hypothalamic, anterior pituitary, and adrenocortical mRNAs was assessed by real-time PCR, and expression of specific proteins in isolated adrenal mitochondria from adrenal zona fascisulata/reticularis was assessed by immunoblot analyses. Oxygen saturation, heart rate, and body temperature were also measured. Exposure to 8% O(2) for as little as 1 h elicited an increase in plasma corticosterone in all age groups studied, with PD2 pups showing the greatest response ( approximately 3 times greater than PD8 pups). Interestingly, the ACTH response to hypoxia was absent in PD2 pups, while plasma ACTH nearly tripled in PD8 pups. Analysis of adrenal mRNA expression revealed a hypoxia-induced increase in Ldlr mRNA at PD2, while both Ldlr and Star mRNA were increased at PD8. Acute hypoxia decreased arterial O(2) saturation (SPo(2)) to approximately 80% and also decreased body temperature by 5-6 degrees C. The hypoxic thermal response may contribute to the ACTH and corticosterone response to decreases in oxygen. The present data describe a developmentally regulated, differential corticosterone response to acute hypoxia, shifting from ACTH independence in early life (PD2) to ACTH dependence less than 1 wk later (PD8).     

Intermittent hypoxia augments carotid body and ventilatory response to hypoxia in neonatal rat pups.

Carotid bodies are functionally immature at birth and exhibit poor sensitivity to hypoxia. Previous studies have shown that continuous hypoxia at birth impairs hypoxic sensing at the carotid body. Intermittent hypoxia (IH) is more frequently experienced in neonatal life. Previous studies on adult animals have shown that IH facilitates hypoxic sensing at the carotid bodies. On the basis of these studies, in the present study we tested the hypothesis that neonatal IH facilitates hypoxic sensing of the carotid body and augments ventilatory response to hypoxia. Experiments were performed on 2-day-old rat pups that were exposed to 16 h of IH soon after the birth. The IH paradigm consisted of 15 s of 5% O2 (nadir) followed by 5 min of 21% O2 (9 episodes/h). In one group of experiments (IH and control, n = 6 pups each), sensory activity was recorded from ex vivo carotid bodies, and in the other (IH and control, n = 7 pups each) ventilation was monitored in unanesthetized pups by plethysmography. In control pups, sensory response of the carotid body was weak and was slow in onset (approximately 100 s). In contrast, carotid body sensory response to hypoxia was greater and the time course of the response was faster (approximately 30 s) in IH compared with control pups. The magnitude of the hypoxic ventilatory response was greater in IH compared with control pups, whereas changes in O2 consumption and CO2 production during hypoxia were comparable between both groups. The magnitude of ventilatory stimulation by hyperoxic hypercapnia (7% CO2-balance O2), however, was the same between both groups of pups. These results demonstrate that neonatal IH facilitates carotid body sensory response to hypoxia and augments hypoxic ventilatory chemoreflex.     

Chronotropic response of cultured neonatal rat ventricular myocytes to short-term fluid shear

Ventricular myocytes are continuously exposed to fluid shear in vivo by relative movement of laminar sheets and adjacent cells. Preliminary observations have shown that neonatal myocytes respond to fluid shear by increasing their beating rate, which could have an arrhythmogenic effect under elevated shear conditions. The objective of this study is to investigate the characteristics of the fluid shear response in cultured myocytes and to study selected potential mechanisms. Cultured neonatal rat ventricular myocytes that were spontaneously beating were subjected to low shear rates (5-50/s) in a fluid flow chamber using standard culture medium. The beating rate was measured from digital microscopic recordings. The myocytes reacted to low shear rates by a graded and reversible increase in their spontaneous beating rate of up to 500%. The response to shear was substantially attenuated in the presence of the beta-adrenergic agonist isoproterenol (by 86+/-8%), as well as after incubation with integrin-blocking RGD peptides (by 92+/-8%). The results suggest that the beta-adrenergic signaling pathway and integrin activation, which are known to interact, may play an important role in the response mechanism.     

Calpastatin is up-regulated in response to hypoxia and is a suicide substrate to calpain after neonatal cerebral hypoxia-ischemia.

In a model of cerebral hypoxia-ischemia in the immature rat, widespread brain injury is produced in the ipsilateral hemisphere, whereas the contralateral hemisphere is left undamaged. Previously, we found that calpains were equally translocated to cellular membranes (a prerequisite for protease activation) in the ipsilateral and contralateral hemispheres. However, activation, as judged by degradation of fodrin, occurred only in the ipsilateral hemisphere. In this study we demonstrate that calpastatin, the specific, endogenous inhibitor protein to calpain, is up-regulated in response to hypoxia and may be responsible for the halted calpain activation in the contralateral hemisphere. Concomitantly, extensive degradation of calpastatin occurred in the ipsilateral hemisphere, as demonstrated by the appearance of a membrane-bound 50-kDa calpastatin breakdown product. The calpastatin breakdown product accumulated in the synaptosomal fraction, displaying a peak 24 h post-insult, but was not detectable in the cytosolic fraction. The degradation of calpastatin was blocked by administration of CX295, a calpain inhibitor, indicating that calpastatin acts as a suicide substrate to calpain during hypoxia-ischemia. In summary, calpastatin was up-regulated in areas that remain undamaged and degraded in areas where excessive activation of calpains and infarction occurs.     

Thymectomy in the neonatal rat.

A method for removal of the thymus in the newborn rat employing measures aimed at reducing the mortality incurred by the surgical procedure, cannibalism and infection is described in detail. Tranquilisation of the dam, sterile technique, magnification of the operative site for thymus aspiration, warming to 37 degrees C during the recovery phase, and application of pheromones to the offspring prior to their reunion with the dam, all proved important in increasing the survival rate.