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Biomedical subjects

R Gyurko

Publications and source records attributed to R Gyurko.

9 recordsLinked to original sources
PubMed1997

Time course of inhibition of hypertension by antisense oligonucleotides targeted to AT1 angiotensin receptor mRNA in spontaneously hypertensive rats.

Antisense oligodeoxynucleotides (AS-ODN) can be designed to provide inhibition of a specific protein. Since angiotensin receptors are involved in blood pressure regulation we constructed AS-ODN to angiotensin II type-1 receptor (AT1) mRNA. When given centrally, the AS-ODN reduces blood pressure in spontaneously hypertensive rats (SHR) 24 h after injection. To study the time course of a single AS-ODN injection on blood pressure and heart rate, groups of SHR were injected intracerebroventricularly (icv) with either single dose of AS-ODN or scrambled (SC) ODN and blood pressure was recorded through implanted catheters daily for up to 9 days. Blood pressure decreased significantly in the AS-ODN treated rats compared to the SC-ODN rats for up to 7 days. The maximum decrease (38 mm Hg) occurred at 3 days. There appeared to be no toxic reaction or side effects and the blood pressure level had recovered by days 8 and 9. Heart rate was not altered by AS-ODN treatment. To test that the ODN was entering the brain tissue, fluorescein-isothiocyanate labelled (FITC) ODN was injected in Sprague-Dawley rats and the fluorescence detected 1 h later by confocal microscopy. Within 1 h there was rapid uptake into cells close to the site of injection and into brain parenchyma around the third and lateral ventricles. To test that the AS-ODN had reduced AT1 receptors, binding studies were carried out on membranes from hypothalamic tissue. There was a modest (approximately 20%) but significant (P < .05) decrease in the AT1 receptor binding after 25 microm or 50 microm AS-ODN. AT2 receptors were not altered by the AS-ODN, indicating its specificity for the AT1 receptor. The small decrease in receptor binding, relative to its large effect on blood pressure, is discussed in terms of the AT1 receptor life cycle. The mechanism for the long action of a single AS-ODN injection is hypothesized as resulting from the persistence of AS-ODN in the nucleus, preventing transport of the mRNA into the cytoplasm.

Animals↗
PubMed1995

A decrease in angiotensin receptor binding in rat brain nuclei by antisense oligonucleotides to the angiotensin AT1 receptor.

Intracerebroventricular (i.c.v.) injections of antisense oligonucleotides against mRNA of the angiotensin type 1 (AT1) receptor have been shown to reduce blood pressure in spontaneously hypertensive (SHR) rats and angiotensin II-induced drinking in both SHR and Sprague-Dawley (SD) rats. The present investigation was designed to quantify the effect of i.c.v. injections of antisense oligonucleotides to the AT1 receptor mRNA on brain angiotensin receptors using membrane binding and autoradiographic analysis. Control injections contained sense or scrambled oligonucleotides or saline. Three daily injections of antisense oligonucleotides into the third ventricle of SD rats decreased the AT1 receptor number significantly by 25% in a hypothalamic tissue block. AT2 receptors were not altered. Autoradiography showed a decrease in angiotensin receptor number in hypothalamic nuclei and in the anteroventral region of the third ventricle (AV3V) after antisense treatment. AT2 receptors were not reduced indicating the AT1 antisense oligonucleotides were specific. In a second series of experiments, single injections of antisense oligonucleotides into the lateral ventricle of SHR rats were tested. Antisense oligonucleotides produced a significant decrease in receptor number in the same hypothalamic area. Sense and scrambled oligonucleotides did not decrease the receptor numbers significantly. The decreases observed after injection of antisense oligonucleotides were between 15 and 30%. These changes may be sufficient to account for the physiological effects of i.c.v. injections of antisense oligonucleotides to AT1 receptor mRNA.

Analysis of Variance↗
PubMed1995

Antisense inhibition of hypertension in the spontaneously hypertensive rat.

Phosphorothioated antisense oligodeoxynucleotide (ASODN) targeted to angiotensinogen mRNA was administered intracerebroventricularly in spontaneously hypertensive rats to test whether angiotensinogen reduction would lower their hypertensive blood pressures. The ASODN lowers hypertensive blood pressures to normotensive levels in spontaneously hypertensive rats; sense oligodeoxynucleotide had no effect. Administration of phosphorothioated ASODN produced a prolonged duration of lowered blood pressure. Injections of ASODN at the same dose that decreased hypertension when administered centrally did not result in blood pressure decreases when administered intra-arterially. Furthermore, angiotensinogen production was decreased in the brain stem and significantly decreased in the hypothalamus of the ASODN-treated rats (P < .05), supporting the concept of centrally mediated regulation of hypertension by an overactive brain angiotensin system. To determine the distribution of centrally administered oligodeoxynucleotides, fluorescein isothiocyanate-conjugated oligodeoxynucleotides were injected directly into the lateral ventricles. One hour later, oligodeoxynucleotides were distributed throughout the lateral and third ventricles, with tissue and cellular uptake observed in discrete cells at the injection site. This indicates that the oligodeoxynucleotides are taken up rapidly by brain cells and that they permeate the areas surrounding brain nuclei involved in central blood pressure regulation and volume homeostasis. The results confirm and extend our previous study with phosphodiester ASODN and show that phosphorothioation modification increases the duration of the response and is taken up in vivo. We conclude that with modification, ASODN inhibition of angiotensinogen mRNA translation can be used for a prolonged, profound decrease in mean arterial pressure in the spontaneously hypertensive rat through a central mechanism.

Angiotensinogen↗
PubMed1994

Antisense oligonucleotide to AT1 receptor mRNA inhibits central angiotensin induced thirst and vasopressin.

Antisense oligodeoxynucleotides (AS-ODN) to AT1 receptor mRNA inhibit high blood pressure in Spontaneously Hypertensive Rats (SHR) when injected into the brain. The effect is presumably through inhibition of the actions of brain angiotensin II (Ang II). Central injection of Ang II elicits several physiological responses including release of vasopressin and motivation to drink. The angiotensin II type-I (AT1) receptor is located in brain regions which have been implicated in mediating these effects. Therefore we hypothesized that AS-ODN to AT1 mRNA would inhibit the drinking and AVP response to central administration of Ang II in adult male SHR. AS-ODN were constructed to bases +63 to +77 (15-mer) of the AT1 receptor RNA. 24 h after AS-ODN treatment (50 micrograms/4 microliters) (intracerebroventricularly, i.c.v.), the drinking response to Ang II (50 ng, i.c.v.) was significantly reduced in the SHR (P < 0.05). The drinking response to Ang II (i.c.v.) was also reduced in the Sprague-Dawley rats (P < 0.05). There was no reduction of water intake in the control animals treated with scrambled ODN (SC-ODN). Repeated injection of AS-ODN did not produce a greater reduction in drinking response. Arginine vasopressin (AVP) release to central Ang II was significantly decreased after AS-ODN treatment when compared to vehicle (P < 0.05) and to SC-ODN injections (P < 0.05). Radioligand binding assays of the hypothalamic block after AS-ODN treatment showed a significant decrease of AT1 receptor binding (P < 0.05). The results show that the antisense inhibition of brain AT1 receptor gene expression decreases the Ang II induced drinking and AVP release responses.

Animals↗
PubMed1994

Antisense inhibition of hypertension: a new strategy for renin-angiotensin candidate genes.

There are several ways of experimentally studying the influence of candidate genes on hypertension. The approach proposed here is antisense inhibition with antisense oligodeoxynucleotides (AS-ODNs) constructed to the 5' region of known sequences of angiotensinogen mRNA and angiotensin II type-1 receptor mRNA. The AS-ODNs were applied in vivo and in vitro. In vivo, direct injection of 50 micrograms of AS-ODN into the lateral ventricles of SHR reduced hypertension significantly (P < 0.01). There was no effect of AS-ODN i.c.v. in normotensive WKY rats. The phosphorothiated AS-ODN to the AT1 receptor mRNA also produced a long-lasting decrease in blood pressure in SHR (7 days). After AS-ODN treatment AT1 receptors were reduced in the PVN and anterior third ventricle area and Ang II levels were reduced in the brainstem. The results show the in vivo feasibility of using antisense inhibition of renin-angiotensin mRNA to reduce hypertension.

Angiotensinogen↗
PubMed1993

Antisense inhibition of AT1 receptor mRNA and angiotensinogen mRNA in the brain of spontaneously hypertensive rats reduces hypertension of neurogenic origin.

To determine the role of angiotensinogen and angiotensin II type-1 (AT1) receptor genes in hypertension, spontaneously hypertensive rats (SHR) were injected with synthetic antisense oligodeoxynucleotides (ODNs), intracerebroventricularly (i.c.v). Antisense ODNs were constructed to bases -5 to +13 of angiotensinogen mRNA (18-mer) and to bases +63 to +77 (15-mer) of angiotensin II type-1 receptor mRNA. Hypertension was significantly reduced by the application of 50 micrograms of both antisense ODNs to normotensive levels. The phosphorothioated antisense ODN to the AT1 receptor produced long-lasting (7 days) decreases in blood pressure. After AT1 antisense treatment, AT1 receptors were reduced in the paraventricular nucleus (PVN) and in the anterior third ventricle area (AV3V). Following angiotensinogen antisense treatment, angiotensin II levels were significantly reduced in the brainstem (P < 0.05), indicating arrest of angiotensin II synthesis. The results demonstrate that inhibiting the brain renin-angiotensin system by antisense inhibition of the angiotensinogen and the AT1 receptor genes, lowers high blood pressure in the SHR. The antisense administration to specific genes of the tissue renin-angiotensin system offers the possibility of a new approach to developing antihypertension treatments.

Angiotensinogen↗
PubMed1992

Angiotensin II receptor subtypes play opposite roles in regulating phosphatidylinositol hydrolysis in rat skin slices.

Among the many functions of angiotensin II (Ang II) it now appears that Ang II is a growth factor. The concentration of Ang II in rat skin has been shown to increase during wound healing. To investigate the intracellular effect of Ang II in skin we determined the levels of total cytoplasmic inositol phosphates after incubation of skin slices with different doses of Ang II. 10(-6) M of Ang II increased significantly the phosphatidylinositol (PI) hydrolysis, and the effect was dose dependent up to 10(-4) M Ang II. The majority of inositol phosphates yielded after 1 hour incubation in the presence of lithium was InsP1, with lesser amount of InsP2. Losartan, the Ang II AT1 antagonist, at a dose of 10(-4) M blocked the effect of Ang II, while PD123319, the Ang II AT2 antagonist, had no antagonistic action; PD123319 at the higher dose of 10(-3) M, however, potentiated the effect of Ang II on PI hydrolysis. The results suggest that PI hydrolysis is a second messenger system for Ang II in rat skin. Also, the two subtypes of Ang II receptors mediate opposite effects on PI hydrolysis: Ang II binding to AT1 receptors increases inositol phosphate production, while Ang II binding to AT2 receptors decreases inositol phosphate production.

Angiotensin II↗