In its classical definition, the renin-angiotensin system (RAS) acts predominantly by endocrine mechanisms. This view has been modified since several components of the RAS and their mRNAs were found ...in peripheral tissues. These findings gave rise to the concept of local tissue renin-angiotensin systems. Although no cells of cardiovascular organs containing a complete RAS have been identified as of this writing, angiotensinogen and angiotensin-converting enzyme (ACE) are most likely synthesized within the vasculature for example. Local synthesis of renin may be limited to very small amounts, but uptake of renin from the circulation is very likely. The function of local RASs is shown by reduction of blood pressure by ACE inhibitors, which correlates better with the inhibition of ACE activity in certain tissues than with its activity in the plasma. Further studies have put forward the notion that the circulating RAS could mainly be important for acute hemodynamic stability, whereas the tissue RAS could be involved in more long term maintenance of hemodynamics. This review will try to summarize the findings leading to the concept of a local tissue renin-angiotensin system, and discuss interactions between the circulating and the local RAS in the light of recent experimental findings.
To evaluate the expression of components of the endothelin (ET) system in single Purkinje neurons and Bergmann glial cells in situ, patch-clamp recording was combined with a multiplex RT-PCR ...approach. Cerebellar slices were rapidly isolated from 20- to 28-day-old mice. Cells were characterized morphologically and electrophysiologically and cell contents were aspirated and immediately reverse-transcribed. The cDNA was used as a template in a multiplex PCR reaction containing primers specific for ET-1, ET-2, and ET-3, ET-converting enzyme 1 (ECE-1) and ECE-2, and ETA and ETB receptors. The resulting PCR products were used as templates in a second PCR reaction containing only one pair of nested primers. Specific single bands were obtained from positive cells, which was confirmed by DNA sequencing of the PCR products. Of the 25 Purkinje neurons assayed, 84% were positive for ECE-1 mRNA and 68% for ECE-2 mRNA. No ET and ETA receptor mRNAs were detected, and only one cell was positive for ETB receptor mRNA. In Bergmann glial cells, ETB receptor mRNA was predominant. A total of 68% of the 25 cells assayed were positive. Sixteen percent were positive for ETA receptor mRNA, 8% for ECE-1 mRNA, and 12% for ECE-2 mRNA. Again, no ET mRNAs were detected. These results confirm the role of the ETB receptor in Bergmann glial cells and provide evidence for expression of ECE-1 and ECE-2 in Purkinje neurons.
Recombinant human erythropoietin (rHuEpo) has been widely used in patients undergoing chronic hemodialysis treatment to correct anemia. In a subgroup of patients, i.v. administration of rHuEpo leads ...to manifestation or worsening of hypertension. The underlying mechanism of this remains unclear but it has been suggested that it is associated with increased expression of the vasoconstrictor endothelin (ET) in endothelial cells (ECs). There is also evidence for expression of specific rHuEpo receptors on ECs. The aim of this work was to study the time course and mechanisms of ET-1 regulation on the mRNA level in bovine aortic endothelial cells (BAECs) and human umbilical vein endothelial cells (HUVECs) stimulated with pharmacologic doses of rHuEpo (1-10 IU/ml). Compared to vehicle-treated controls, rHuEpo-treatment of ECs increases preproET-1 mRNA expression up to 170%, as shown by Northern blotting. To study the transcriptional regulation of ET-1 expression by rHuEpo, ECs were transfected with a luciferase construct driven by the rat ET-1 promoter and subsequently stimulated with rHuEpo. Compared to controls, luciferase activity increased up to 200% (n = 6; p < 0.05), suggesting transcriptional regulation of preproET-1 mRNA-expression by rHuEpo. Our data support the hypothesis that ET contributes to the hypertensive side effects of rHuEpo treatment and that this interaction occurs at the transcriptional level.
The local effects of angiotensin II (ANG II) on the heart may play an important role for the pathophysiology of cardiovascular disease. Numerous in vitro studies have demonstrated that angiotensin II ...has distinctive cellular effects in the cardiovascular system which are independent from its effects on blood pressure. These have led to the hypothesis that activation of the angiotensin system in the heart could be of functional relevance for the adaptive processes in several cardiovascular disorders such as cardiac hypertrophy heart failure. This concept has been further supported by clinical studies showing the beneficial effects of angiotensin-converting enzyme inhibitors in these circumstances. In order to study the gene regulation of renin-angiotensin system components in cardiac disorders we investigated the gene expression of angiotensin converting enzyme in human heart failure. Results showed that the enzyme is activated locally in this condition, supporting previous studies in animals. Taken together with recent evidence from genetic studies linking the enzyme to myocardial infarction and cardiac hypertrophy, our findings are in support of the notion that angiotensin converting enzyme plays a central role in cardiovascular physiology and pathophysiology.
The endothelin (ET) isoforms ET-1, ET-2 and ET-3 applied at 100 nM triggered a transient increase in Ca
2+
i in Bergmann glial cells in cerebellar slices acutely isolated from 20–25 day-old mice. The ...intracellular calcium concentration (Ca
2+
i) was monitored using Fura-2-based (Ca
2+
i) microfluorimetry. The ET-triggered (Ca
2+
i) transients were mimicked by ET, receptor agonist BO-3020 and were inhibited by ETB receptor antagonist BQ-788. ET elevated Ca
2+
i in Ca2+-free extracellular solution and the ET-triggered Ca
2+
i elevation was blocked by 500 nM thapsigargin indicating that the Ca
2+
i was released from InsP3 sensitive intracellular pools. The ET-triggered Ca
2+
i increase in Ca
2+-free solution was shorter in duration. Restoration of normal extracellular Ca
2+ briefly after the ET application induced a second Ca
2+
i increase indicating the presence of a secondary Ca
2+ influx which prolongs the Ca
2+ signal. Pre-application of 100 μM ATP or 10 μM noradrenaline blocked the ET response suggesting the involvement of a common Ca
2+ depot. The expression of ETB receptor mRNAs in Bergmann glial cells was revealed by single-cell RT-PCR. The mRNA was also found in Purkinje neurones, but no Ca
2+ signalling was triggered by ET. We conclude that Bergmann glial cells are endowed with functional ET
B receptors which induce the generation of intracellular Ca
2+
i signals by activation of Ca
2+ release from InsP
3-sensitive intracellular stores followed by a secondary Ca
2+ influx.
The endothelin (ET) isoforms ET-1, ET-2 and ET-3 applied at 100 nM triggered a transient increase in Ca2+i in Bergmann glial cells in cerebellar slices acutely isolated from 20-25 day-old mice. The ...intracellular calcium concentration (Ca2+i) was monitored using Fura-2-based Ca2+i microfluorimetry. The ET-triggered Ca2+i transients were mimicked by ETB receptor agonist BQ-3020 and were inhibited by ETB receptor antagonist BQ-788. ET elevated Ca2+i in Ca(2+)-free extracellular solution and the ET-triggered Ca2+i elevation was blocked by 500 nM thapsigargin indicating that the Ca2+i was released from InsP3-sensitive intracellular pools. The ET-triggered Ca2+i increase in Ca(2+)-free solution was shorter in duration. Restoration of normal extracellular Ca2+ briefly after the ET application induced a second Ca2+i increase indicating the presence of a secondary Ca2+ influx which prolongs the Ca2+ signal. Pre-application of 100 microM ATP or 10 microM noradrenaline blocked the ET response suggesting the involvement of a common Ca2+ depot. The expression of ETB receptor mRNAs in Bergmann glial cells was revealed by single-cell RT-PCR. The mRNA was also found in Purkinje neurones, but no Ca2+ signalling was triggered by ET. We conclude that Bergmann glial cells are endowed with functional ETB receptors which induce the generation of intracellular Ca2+i signals by activation of Ca2+ release from InsP3-sensitive intracellular stores followed by a secondary Ca2+ influx.
