Actions of cyclosporin A on cultured rat mesangial cells

The effects of cyclosporin A (CsA) on planar surface area of cultured rat mesangial cells (PCSA) and cross-sectional area of isolated rat glomeruli (GCSA) were tested. The same experiments were performed after preincubation with platelet activating factor (PAF) antagonists (BN 52021, alprazolam) or calcium channel blockers (verapamil). Areas of cells or glomeruli were analyzed by a computer-assisted method. CsA reduced PCSA in a time-dependent (significant effects began at 15 min, about 12% of reduction with 10(-6) M CsA) and dose-dependent (no effect at 10(-9) M CsA, maximal reduction of about 30% at 60 min of incubation with 10(-7) M CsA) fashion. BN 52021 (5.10(-5) M) and alprazolam (10(-5) M) completely inhibited the reduction of mesangial cell area induced by CsA. Verapamil (10(-5) M) only partially inhibited this action. Glomerular cross-sectional area decreased after 30 minutes of incubation with 10(-6) M CsA (1.45 +/- 0.02 vs. 1.55 +/- 0.02 m2.10(-8), P less than 0.001), an effect that was inhibited by BN 52021 or verapamil. In addition, 10(-6) M CsA increased PAF production by isolated rat glomeruli (425 +/- 80 pg/mg vs. 198 +/- 13 pg/mg in control glomeruli, P less than 0.01), an effect which was not inhibited by verapamil. These results suggest that CsA could reduce GFR by decreasing the glomerular ultrafiltration coefficient, perhaps as a consequence of the contraction of mesangial cells. PAF seems to play a pivotal role in the genesis of this action.     

Role of cGMP as second messenger of adenosine in the inhibition of renin release

Adenosine is known to be a potent inhibitor of renin release from the kidneys. The aim of this study was to investigate the transmembrane signalling avenue that the second messenger of adenosine causes inhibition of renal renin release. Using short term cultures of juxtaglomerular cells isolated from rat kidneys, we found that adenosine inhibited spontaneous renin release from these cells up to 40% of control, in a dose dependent fashion between 10(-10) M to 10(-6) M. Half maximal inhibition was observed at 2 X 10(-8) M adenosine. The inhibitory effect of adenosine on renin release could be mimicked by the A1-receptor agonist N6-cyclohexyladenosine (CHA) and could be attenuated by the A-receptor antagonist theophylline (5 X 10(-5) M). The A2-receptor agonist 5'-N-ethylcarboxamideadenosine (NECA) had no inhibitory effect on renin release. These findings indicate that the inhibitory effect of adenosine is mediated by A1-receptors on juxtaglomerular cells. Adenosine had no effect on either transmembrane calcium influx or the cytosolic free calcium concentration in the isolated juxtaglomerular cells. Adenosine also did not alter the cellular level of cyclic AMP in the concentration range employed. However, adenosine led to a dose dependent increase of the cellular level of cyclic GMP. Half maximal increase of cGMP was observed at 10(-8) M adenosine. The effect of adenosine on cyclic GMP could be mimicked by the A1-receptor agonist CHA and could be attenuated by the A-receptor antagonist, theophylline.(ABSTRACT TRUNCATED AT 250 WORDS)     

Preconditioning and adenosine protect human proximal tubule cells in an in vitro model of ischemic injury

Renal ischemic reperfusion injury results in unacceptably high mortality and morbidity during the perioperative period. It has been recently demonstrated that ischemic preconditioning or adenosine receptor modulations attenuate renal ischemic reperfusion injury in vivo. An in vitro model of ischemic renal injury was used in cultured human proximal tubule (HK-2) cells to further elucidate the protective signaling cascades against renal ischemic reperfusion injury. ATP depletion preconditioning (1 h of antimycin A and 2-deoxyglucose treatment followed by 1 h of recovery), adenosine, an A(1) adenosine receptor selective agonist, or an A(2a) adenosine receptor selective agonist significantly attenuated subsequent severe ATP depletion injury of HK-2 cells. In contrast, an adenosine receptor antagonist failed to prevent protection induced by ATP depletion preconditioning. Cytoprotection by ATP depletion preconditioning or A(1) adenosine receptor activation was prevented by inhibitors of extracellular signal-regulated mitogen-activated kinases, protein kinase C, and tyrosine kinases. The A(1) and A(2a) adenosine receptor-mediated cytoprotection were also dependent on G(i/o) proteins and PKA activation, respectively. It is concluded that ATP depletion preconditioning and A(1) and A(2a) adenosine receptor activation protect HK-2 cells against severe ATP depletion injury via distinct signaling pathways.     

Leukotriene C4 and D4 contract rat glomerular mesangial cells

The sulfidopeptide leukotrienes, LTC4 and LTD4, have vasoconstrictor effects in the kidney, reducing both renal blood flow and the glomerular filtration rate. As one mechanism regulating the glomerular filtration rate, mesangial cell contraction may reduce the capillary surface area, thereby lowering the ultrafiltration coefficient. Using image analysis microscopy to quantify changes in cell morphology, we found that LTC4 and LTD4 (1 X 10(-12)M to 1 X 10(-6)M) reduced the cross-sectional area of cultured mesangial cells from rat glomeruli. The response to LTC4 and LTD4 (10(-6)M), as measured by the percentage of responding cells (30 to 35%), the maximum decrease in cross-sectional area (25 to 32%), and the time course was identical to that for angiotensin II (10(-6)M). The contraction induced by LTD4 was attenuated by an LTD4 receptor antagonist (4R,5S,6Z-nor-LTD1). Also, preincubation with colchicine prevented LTC4-induced contraction. Leukotriene B4, a non-sulfidopeptide leukotriene that stimulates chemotaxis and chemokinesis, had negligible agonist activity. Mesangial cells cultured on less adhesive teflon membranes were more responsive to LTD4 (62% of cells responded) than cells cultured on glass or polystyrene (35% of cells responded). Mesangial cell contraction was not merely a shape-change as a result of cell damage, since cellular injury was not documented by lactate dehydrogenase release and proliferation of mesangial cells was not retarded by LTC4. Furthermore, the contraction was independent of cell size. Because leukotrienes stimulate cyclooxygenase products in other cells, we examined the ability of the sulfidopeptide leukotrienes to stimulate prostaglandin and thromboxane synthesis. LTC4 and LTD4 did not stimulate PGE2 formation, the major cyclooxygenase product of rat mesangial cells.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of adenosine on isolated afferent arterioles

We investigated the direct effect of adenosine on afferent arterioles (Af-Arts) and the receptor subtype that mediates the constrictor or dilator action of adenosine. Af-Arts were isolated from the superficial cortex of rabbit kidney and perfused in vitro. Adenosine added to either the lumen or bath constricted the Af-Arts in a dose-dependent manner. This constriction was blocked by the A1 receptor antagonist, 6-oxo-3-(2-phenylpyrazole(1,5-a)pyridin-3-yl)-1 (6H)-pyridazinebutyric acid (FK838) or 8-cyclopentyl-1, 3-dipropylxanthine(DPCPX). We also examined the effect of adenosine on preconstricted Af-Arts with norepinephrine. Adenosine added to either the lumen or bath further constricted the preconstricted Af-Arts. In the presence of FK838, adenosine added to either the lumen or bath dilated the preconstricted Af-Arts, but in a different dose-dependent manner. Adenosine-induced dilation was inhibited by the A2 receptor antagonist, 3, 7-dimetyl-1-propargylxanthine(DMPX). These data indicate that adenosine constricts Af-Arts via A1 receptors and that adenosine dilates preconstricted Af-Arts via A2 receptors when A1 receptors are blocked.     

The mechanism of contraction by 2-chloroadenosine in cat detrusor muscle cells

PURPOSE: Four adenosine receptors (ARs), designated A1AR (A1 adenosine receptor), A2aAR (A2a adenosine receptor), A2bAR (A2b adenosine receptor), and A3AR (A3 adenosine receptor), have been cloned from various species, but the contraction mechanism via A1ARs in cat detrusor muscle cell is not well known. MATERIALS AND METHODS: We examined the cellular mechanism using an A1AR agonist 2-chloroadenosine (2-CA) in cat detrusor cell isolated by enzymatic digestion. To examine which phospholipase mediates the contraction, we used phospholipase inhibitors. RESULTS: The adenosine analog potency order is R-N6-phenylisopropyladenosine (R-PIA) > 5'-N-ethylcarbosamine adenosine (NECA) > 2-chloroadenosine (2-CA) > S-N6-phenylisopropyladenosine (S-PIA). The ratio of equi-effective concentrations of R-PIA/S-PIA was 58.2. 8-cyclopentyl-1,3-dipropylxanthine (DPCPX, 300 nM) shifted to the right the concentration-response curves of 2-CA. These results indicate A1ARs mediate 2-CA induced contraction in cat detrusor muscle. G proteins (Gi1, Gi2, Gi3, Go, Gs, and Gq) in cat detrusor muscle were detected by immunoblot analysis. Pertussis toxin (PTX) inhibited 2-CA induced contraction. In permeabilized cells, antibodies against Galphai3 antagonized 2-CA induced contraction, suggesting that the contraction is mediated by Gi3 protein. A phosphatidylinositol-specific phospholipase C (PLC) inhibitor, neomycin, reduced 2-CA induced contraction, but a phospholipase D (PLD) inhibitor, p-chloromercuribenzoic acid, and a phospholipase A2 (PLA2) inhibitor, dimethyl-eicosa-dienoic acid (DEDA), had no effect. We found the presence of the main PLC isozymes, PLC-beta1, PLC-beta3, and PLC-gamma1. 2-CA induced contraction in permeabilized cells was inhibited by PLC-beta3 but not by PLC-beta1 or PLC-gamma1 antibody. These results imply that A1ARs are coupled to PLC-beta3 via PTX-sensitive Gi3 protein. Sr2+ medium and thapsigargin, which replaces intracellular Ca2+ and deplete intracellular calcium stores respectively, inhibited 2-CA induced contraction. CONCLUSIONS: These results suggest that A1ARs mediating 2-CA induced contraction exist in cat detrusor muscle and the contraction depends on a PTX-sensitive Gi3 protein, PLC-beta3 and the release of intracellular Ca2+.     

Adenosine is upregulated during peritonitis and is involved in downregulation of inflammation

Loss of function of the peritoneal membrane is associated with peritonitis. Adenosine levels in sites of inflammation were shown to increase and exhibit immunoregulatory effects. Our aim was to elucidate the regulatory role of adenosine during peritonitis and to test the involvement of peritoneal mesothelial cells (PMC) in adenosine regulation. In a mice model of Escherichia coli peritonitis, the adenosine A(2A) receptor (A(2A)R) agonist (CGS21680) prevented leukocyte recruitment and reduced tumor necrosis factor alpha (TNF-alpha) and interleukin-6 (IL-6) levels. Peritonitis induced the elevation of adenosine with a peak at 24 h. Analysis of adenosine receptor levels on peritoneum showed that A(1) receptor (A(1)R) protein levels peak at 12 h after inoculation and then return to baseline at 24 h, whereas high affinity A(2A)R protein levels peak at 24 h concomitantly with the peak of adenosine concentration. Low affinity A(2B) receptor (A(2B)R) levels elevated slowly, remaining elevated up to 48 h. In human PMC (HPMC), the early cytokines, IL-1-alpha, and TNF-alpha upregulated the A(2B) and A(2A) receptors. However, interferon-gamma (IFN-gamma) upregulated the A(2B)R and decreased A(2A)R levels. Treatment with the A(2A)R agonist reduced IL-1-dependent IL-6 secretion from HPMC. In conclusion, the kinetics of adenosine receptors suggest that at early stage of peritonitis, the A(1)R dominates, and later its dominance is replaced by the G stimulatory (Gs) protein-coupled A(2A)R that suppresses inflammation. Early proinflammatory cytokines are an inducer of the A(2A)R and this receptor reduces their production and leukocyte recruitment. Future treatment with adenosine agonists should be considered for attenuating the damage to mesothelium during the course of acute peritonitis.     

Functional prostaglandin E (EP) receptors in human penile corpus cavernosum

In this study, we have characterized functional EP receptors in human corpus cavernosum (HCC) tissue and in HCC smooth muscle cells (SMC). Using RNase protection assays, we determined expression of EP2, EP3I and EP3II receptor mRNA. In organ bath preparations of HCC tissue strips, PGE1 caused dose-dependent relaxation at concentrations below 300 nM. At concentrations greater than 300 nM, PGE1 caused contraction. Addition of the EP1/EP2/EP3 receptor antagonist AH6809 inhibited this contraction and facilitated further relaxation through concentrations above 1 microM of PGE1. The EP1/EP3 receptor selective agonist 17-phenyltrinor-PGE2 caused dose-dependent contraction that was partially attenuated by SC51322, an EP1 selective antagonist. In cultures of HCC SMC, PGE1 stimulated cAMP accumulation in a dose-dependent manner. Interestingly, AH6809 significantly attenuated PGE1-induced cAMP accumulation. Sulprostone, a selective EP3 receptor agonist, induced weak contractions in HCC tissue strips but augmented forskolin-induced cAMP synthesis in HCC SMC. The data in this study suggest that HCC and cultured smooth muscle cells express EP1, EP2 and EP3 receptors. These receptors mediate their responses via different biochemical pathways and are expected to have different responses in regulating smooth muscle tone. Thus, we suggest that the ultimate response in erectile tissue to various prostanoids is the integration of responses elicited by individual EP receptor subtypes to a given ligand.     

Calcium-mobilizing purine receptors on the surface of mammalian articular chondrocytes

If we are to fully understand mechanisms of cartilage homeostasis, it is essential that we know the full catalogue of receptors present on the surface of a chondrocyte and the pathways regulated by ligands that bind to these receptors. In this study, we describe chondrocyte responses to adenosine 5′-triphosphate and related molecules. Adenosine 5′-triphosphate stimulated a statistically significant, dose-dependent, transient rise in the concentration of calcium ions in Fura 2-loaded, differentiated, primary chondrocytes. The increase occurred in the absence of extracellular calcium, indicating a mobilization from intracellular stores. The increase in concentration of cytoplasmic calcium ions induced by adenosine 5′-triphosphate was mimicked by uridine 5′-triphosphate but not by 2-methylthioadenosine 5′-triphosphate. Cytidine 5′-triphosphate, or adenosine. Heteralogous desensitization experiments demonstrated that chondrocytes showed no subsequent response to uridirie 5′-triphosphate after initial stimulation with adenosine 5′-triphosphate nor did they respond to adenosine 5′triphosphate in inverse conditions, thereby indicating competition for the same receptor site. Together, these results are consistent with the presence of a P2U: receptor on the cell surface of chondrocytes. Purine-induced calcium mobilization in passaged chondrocytes showed the same pharmacological profile with respect to aganist sensitivity, but responses were of greater magnitude than responses in primary differentiated chondrocytes, suggesting upregulation of the receptor with time in culture. Adenosine 5′-triphosphate and uridine 5′-triphosphate (1-100 μM) did not alter cartilage matrix synthesis as measured by rate of incorporation of [³⁵S]sulfate into glycosaminoglycan by cartilage explants or primary chondrocytes. Matrix degradation, measured by release of glycosaminoglycan from cartilage explants, was also unaltered by adenosine 5′-triphosphate or uridine 5′-triphosphate (1-100 μM). Production of prostaglandin E₂ was upregulated by incubation with either adenosine 5′-triphosphate or uridine 5′-triphosphate. These data demonstrate the presence of a functional P_2U,-like purine receptor on the surface of primary articular chondrocytes and support the hypothesis that altered concentrations of extracellular purines may influence chondrocyte metabolism.     

Characterization of myosin heavy and light chains in cultured mesangial cells.

Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) demonstrated that postconfluent mesangial cells in primary culture expressed three myosin heavy chains (MHCs), 204 kD, 200 kD and 196 kD, in a manner similar to that of smooth muscle cells. The MHCs of 204 kD and 200 kD in mesangial cells reacted positively with antibodies raised against bovine aorta smooth muscle myosin while the 196 kD MHC reacted positively with antibodies against platelet myosin. Moreover, the combined content of the MHCs in cultured mesangial cells was remarkably similar in amount to that in cultured aortic smooth muscle cells. After three passages, cultured mesangial cells expressed only the 196 kD MHC as has been reported for cultured smooth muscle cells. Two phosphorylated proteins were found in the immunoprecipitate after incubation of the cell extract with antibodies against platelet myosin: a MHC of approximately 200 kD and myosin light chain (MLC) of 20 kD. The level of MLC phosphorylation was quantitated by scanning densitometry of autoradiograms. Arginine vasopressin (AVP) at 100 nM induced MLC phosphorylation with a maximum effect at 10 minutes. AVP enhanced MLC phosphorylation in a dose dependent manner: maximum response was observed with 100 nM and half maximum, at 3.5 nM. Similarly, angiotensin II (100 nM), endothelin-1 (10 nM) and the calcium ionophore, A23187 (1 microM), significantly enhanced MLC phosphorylation. Thus, although the expression of MHC was altered in quality after mesangial cells were placed in culture, the cells remained rich in myosin content and had an intact regulatory system for contraction which responded to a variety of vasoconstrictive agents.(ABSTRACT TRUNCATED AT 250 WORDS)     

Role of coagulation factor Xa and protease-activated receptor 2 in human mesangial cell proliferation

BACKGROUND: Fibrin deposition and mesangial cell proliferation are frequently observed in the active type of mesangioproliferative glomerulonephritis. Coagulation factors, such as factor V and factor Xa are colocalized with fibrin in the mesangial areas in active type of IgA nephropathy with mesangial cell proliferation. In this study, therefore, we studied the role of factor Xa and its receptor, protease-activated receptor 2 (PAR2) in mesangial cell proliferation and fibrin deposition, and examined ant-proliferative effects of a specific factor Xa inhibitor, DX-9065a, in cultured human mesangial cells. METHODS: To examine the effect of DX-9065a on the factor Xa-induced proliferation of cultured human mesangial cells, we measured thymidine incorporation and cell numbers. We also examined the effect of DX-9065a on extracellular regulated kinase (ERK) activation and fibrin production induced by factor Xa in human mesangial cells. RESULTS: Factor Xa increased [(3)H]-thymidine incorporation and cell numbers in a dose-dependent manner in mesangial cells, which was inhibited by DX-9065a. DX-9065a also suppressed factor Xa-triggered fibrin deposition on mesangial cell surface. Factor Xa induced the activation of ERK in mesangial cells and this activation was also completely inhibited by DX-9065a, but not inhibited by PAR1 antagonist. Factor Xa-induced cell proliferation and ERK activation were inhibited by PD98059. CONCLUSION: There results suggest that factor Xa can induce mesangial cell proliferation through the activation of ERK via PAR2 in mesangial cells and that PAR2 may play a crucial role in the cell proliferation induced by factor Xa. Our results implicate that DX-9065a may be a promising agent to regulate proliferation of mesangial cellss and inhibit the coagulation process in mesangium.     

腺苷A2型受体在肝窦内皮细胞一氧化氮生成中的作用

目的研究腺苷A2型受体在肝窦内皮细胞(HSECs)一氧化氮生成中的作用。方法分别将腺苷、腺苷A1型受体激动剂R鄄PIA和腺苷A2型受体激动剂CGS21680与原代培养的HSEC共培养20min、40min、60min后,用RT鄄PCR测定HSECeNOSmRNA的表达,并测定HSEC内一氧化氮(NO)含量的变化。结果与对照组相比,腺苷和CGS21680都能促使HSEC内eNOSmRNA表达增加,提高HSEC内NO含量(P<0.05),而R鄄PIA却无此作用(P>0.05)。结论HSEC通过腺苷A2型受体的兴奋作用产生大量NO。     

The anticonvulsant action of propofol on epileptiform activity in rat hippocampal slices

We used extracellular electrophysiological recordings from the CA1 region in rat hippocampal slices to investigate the effects of propofol on the field excitatory postsynaptic potential (fEPSP), population spike, and epileptiform activity induced by a Mg(2+)-free condition. Propofol depressed the population spike, fEPSP, and epileptiform activity. Both aminophylline, a nonselective adenosine receptor antagonist, and 8-cyclopentyl-1,3-dipropylxanthine, an A(1) receptor antagonist, significantly reduced the effect of propofol on fEPSP amplitude. However, 3,7-dimethyl-1-propagylxanthine, an A(2) receptor antagonist, did not alter the effect of propofol on fEPSP amplitude. Picrotoxin, a specific chloride channel blocker, partly reduced the effect of propofol on epileptiform activity, but bicuculline, a competitive gamma-aminobutyric acid(A) receptor antagonist, failed to antagonize it. Aminophylline significantly reduced the action of propofol on the epileptiform activity. The anticonvulsant action of propofol was partly reduced by 8-cyclopentyl-1,3-dipropylxanthine, whereas 3,7-dimethyl-1-propagylxanthine failed to affect it. Adenosine depressed the amplitude of fEPSPs in a dose-dependent manner, and propofol enhanced this inhibition. The results demonstrated that, in rat hippocampal slices, propofol inhibits epileptiform activity. In addition, adenosine neuromodulation through the A(1) receptor may contribute to the anticonvulsant action of propofol.     

Effect of calcium channel blockers, nifedipine and benidipine, on death of cultured mouse mesangi al cells

We examined the effects of the short-acting calcium channel blocker (CCB) nifedipine and the long-acting CCB benidipine on the death of mouse cultured mesangial cells induced by tumor necrosis factor alpha (TNF-alpha) and/or cycloheximide (CHX). Cell death was evaluated by a morphological study using semithin sections. The dead cells were divided into three types, i.e., apoptotic cells (type 1), necrotic cells (type 3) and other types of dead cells, the so-called 'secondary necrotic cells' or 'postapoptotic necrotic cells' (type 2). In the morphological study with semithin sections, cells in the presence of TNF-alpha or CHX and nifedipine or benidipine showed low percentages of all dead cell types with 24 h incubation. Both nifedipine and benidipine have protective effects against TNF-alpha or CHX. It is postulated that CCB might inhibit the apoptotic or necrotic processes by TNF-alpha or CHX with 24 h incubation. With 36 h incubation, CCB increased the percentages of all types of dead cells except for treatment with 1x10(-5) M benidipine and CHX. It appears that these cell-protective effects might be decreased after treatment with TNF-alpha or CHX and CCB for 36 h. In conclusion, the short-acting CCB nifedipine and the long-acting CCB benidipine have protective effects on mouse cultured mesangial cells against TNF-alpha or CHX. However, nifedipine and benidipine did not inhibit specific types of cell death using semithin sections in this study.     

Alpha1beta1 integrin-mediated collagen matrix remodeling by rat mesangial cells is differentially regulated by transforming growth factor-beta and platelet-derived growth factor-BB

Pathologic remodeling of mesangial matrix after glomerular injury is the central biologic feature of glomerular scarring (sclerosis). Transforming growth factor-beta (TGF-beta) and platelet-derived growth factor (PDGF)-BB have been implicated in the development of glomerular scarring in rat and human glomerulonephritis. To clarify molecular and cellular mechanisms involved in abnormal mesangial remodeling, this study focused on the role of alpha1beta1 integrin, a collagen/laminin receptor, in rat mesangial cells, using collagen gel contraction as an experimental model of in vivo collagen matrix remodeling and scar formation. In addition, the influence of TGF-beta and PDGF-BB on mesangial cell (MC)-mediated collagen gel contraction in association with the alpha1beta1 integrin expression was evaluated. Integrin function blocking studies using anti-alpha1, beta1 subunit antibodies indicated that MC-alpha1beta1 integrin is essentially required not only for collagen-dependent adhesion/migration, but also for gel contraction. Protein synthesis and mRNA analysis experiments demonstrated that TGF-beta, but not PDGF-BB, increases the expression of alpha1beta1 integrin in mesangial cells cultured on plastic surface and in collagen gels. The upregulation of alpha1beta1 integrin expression by TGF-beta correlated with increases in gel contraction and collagen-dependent adhesion but not migration of mesangial cells. On the other hand, PDGF-BB enhanced MC-mediated gel contraction and migration without affecting cell adhesion to collagen I. Growth factor-induced collagen-dependent adhesion, migration, and gel contraction were significantly attenuated by incubation with anti-alpha1, beta1 subunit antibodies. Thus, these data indicate that alpha1beta1 integrin-mediated collagen matrix remodeling can be modulated by TGF-beta and PDGF-BB via different mechanisms. Alpha1 integrin-mediated mesangial matrix remodeling induced by TGF-beta or PDGF-BB may be a pathogenic mechanism leading to glomerular scarring.     

Adenosine and the Stunned Heart

Abstract Adenosine is one agent under investigation as a therapeutic intervention of myocardial stunning. Adenosine caused numerous effects on the cardiovascular system through its interaction with A₁ and A₂ receptors. We investigated adenosine A₁ receptor mediated mechanisms of cardiac protection in the stunned rat myocardium. Previous studies showed that both adenosine and R-phenylisopropyladenosine (PIA), an A₁ receptor agonist, prolonged the time to onset of ischemic contracture in ischemic isolated rat hearts. Phenylaminoadenosine, an A₂ receptor agonist, did not have any effect, while receptor antagonists blocked adenosine and PIA action. Direct attenuation of the effects of myocardial stunning was observed by altering levels of interstitial fluid adenosine. Our laboratory has shown that administration of erthro-9(2-hydroxy-3-nonyl) adenine (EHNA; an adenosine deaminase inhibitor) to dogs subjected to left anterior descending coronary artery (LAD) occlusion followed by reperfusion results in dramatic increases in ischemic levels of interstitial fluid adenosine and postischemic myocardial function. Using a similar model in dogs, we have shown that exogenous intracoronary adenosine (50 μg/kg per min) augmented postischemic recovery of function, as assessed by significant enhancement (p < 0.01) of systolic wall thickness (7.0 ± 3.0 pretreatment vs ?5.7 ± 1.7 controls). These data support the role for an adenosine A₁ receptor mediated mechanism for protection against myocardial stunning.     

Endothelin and eicosanoid synthesis in cultured mesangial cells

We investigated the effect of endothelin on the generation of eicosanoids, which are known to regulate basal and stimulated mesangial cell tone. The results showed that endothelin is a potent stimulus of prostaglandin E2 (PGE2), prostacyclin (PGI2), and thromboxane A2 (TxA2) synthesis by bovine mesangial cells. Percentage increases in eicosanoid synthesis induced by endothelin (10(-10) to 10(-6) M), were 50 to 275% for PGE2, 28 to 168% for PGI2 and 42 to 111% for TxA2, respectively. Endothelin-induced eicosanoid synthesis in mesangial cells was concentration, but not time dependent. Aspirin (500 microM) completely prevented endothelin-induced eicosanoid synthesis. The calcium entry blocker nitrendipine (10(-8) M) failed to inhibit endothelin-induced eicosanoid synthesis. These data suggest that endothelin-induced changes in renal circulation and glomerular function in normal and disease conditions may be modulated by the concomitant stimulation of mesangial eicosanoid synthesis.