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Biomedical subjects

J Ohanian

Publications and source records attributed to J Ohanian.

At least 19 recordsLinked to original sources
PubMed2001

Diacylglycerol kinase theta is translocated and phosphoinositide 3-kinase-dependently activated by noradrenaline but not angiotensin II in intact small arteries.

Diacylglycerol (DG) kinase (DGK) phosphorylates the lipid second messenger DG to phosphatidic acid. We reported previously that noradrenaline (NA), but not angiotensin II (AII), increases membrane-associated DGK activity in rat small arteries [Ohanian and Heagerty (1994) Biochem. J. 300, 51-56]. Here, we have identified this DGK activity as DGKtheta, present in both smooth muscle and endothelial cells of these small vessels. Subcellular fractionation of artery homogenates revealed that DGKtheta was present in nuclear, plasma membrane (and/or Golgi) and cytosolic fractions. Upon NA stimulation, DGKtheta translocated towards the membrane and cytosol (155 and 153% increases relative to the control, respectively) at 30 s, followed by a return to near-basal levels at 5 min; AII was without effect. Translocation to the membrane was to both Triton-soluble and -insoluble fractions. NA, but not AII, transiently increased DGKtheta activity in immunoprecipitates (126% at 60 s). Membrane translocation and DGKtheta activation were regulated differently: NA-induced DGKtheta activation, but not translocation, was dependent on transient activation of phosphoinositide 3-kinase (PI 3-K). In addition, DGK activity co-immunoprecipitated with protein kinase B, a downstream effector of PI 3-K, and was increased greatly by NA stimulation. The rapid and agonist-specific activation of DGKtheta suggests that this pathway may have a physiological role in vascular smooth-muscle responses.

Androstadienes↗
PubMed2001

Sphingolipids in mammalian cell signalling.

Sphingolipids and their metabolites, ceramide, sphingosine and sphingosine-1-phosphate, are involved in a variety of cellular processes including differentiation, cellular senescence, apoptosis and proliferation. Ceramide is the main second messenger, and is produced by sphingomyelinase-induced hydrolysis of sphingomyelin and by de novo synthesis. Many stimuli, e. g. growth factors, cytokines, G protein-coupled receptor agonists and stress (UV irradiation) increase cellular ceramide levels. Sphingomyelin in the plasma membrane is located primarily in the outer (extracellular) leaflet of the bilayer, whilst sphingomyelinases are found at the inner (cytosolic) face and within lysosomes/endosomes. Such cellular compartmentalisation restricts the site of ceramide production and subsequent interaction with target proteins. Glycosphingolipids and sphingomyelin together with cholesterol are major components of specialised membrane microdomains known as lipid rafts, which are involved in receptor aggregation and immune responses. Many signalling molecules, for example Src family tyrosine kinases and glycosylinositolphosphate-anchored proteins, are associated with rafts, and disruption of these domains affects cellular responses such as apoptosis. Sphingosine and sphingosine-1-phosphate derived from ceramide are also signalling molecules. In particular, sphingosine-1-phosphate is involved in proliferation, differentiation and apoptosis. Sphingosine-1-phosphate can act both extracellularly through endothelial-differentiating gene (EDG) family G protein-coupled receptors and intracellularly through direct interactions with target proteins. The importance of sphingolipid signalling in cardiovascular development has been reinforced by recent reports implicating EDG receptors in the regulation of embryonic cardiac and vascular morphogenesis.

Animals↗
PubMed2001

Lipid second messenger regulation: the role of diacylglycerol kinases and their relevance to hypertension.

Extracellular stimuli elicit cellular responses through generation of intracellular second messengers. The lipid second messenger diacylglycerol is produced following activation of the phosphoinositide signalling system. Diacylglycerol is the physiological activator of protein kinase C but also interacts indirectly with other signalling molecules such as small G proteins. Diacylglycerol kinases convert diacylglycerol to phosphatidic acid so terminating signalling through diacylglycerol. However, phosphatidic acid itself has a lipid second messenger role, with targets distinct from those of its precursor diacylglycerol. Therefore, diacylglycerol kinases occupy a central position in signal transduction and regulation of their activity is crucial to cellular function. A family of nine mammalian diacylglycerol kinases have been identified. Their structural diversity and complex pattern of tissue expression suggests that they function in distinct cellular processes. In addition to the plasma membrane, diacylglycerol kinases are found at the nucleus and cytoskeleton and translocation between subcellular compartments occurs with agonist stimulation. In small arteries diacylglycerol kinase activity is increased by adrenergic stimulation implying a role in vascular smooth muscle responses. Due to their role as key regulators of protein kinase C activity diacylglycerol kinases may play a role in the cardiovascular changes that occur in hypertension and as such could represent novel therapeutic targets.

Diacylglycerol Kinase↗
PubMed2001

Activation of p38 mitogen-activated protein kinases by endothelin and noradrenaline in small arteries, regulation by calcium influx and tyrosine kinases, and their role in contraction.

Small-artery responses to vasoconstrictor agonists are important for vascular function. To investigate the signaling pathways involved in contraction, we studied the activation and regulation of p38 mitogen-activated protein kinases (p38MAPKs) and heat shock protein (HSP) kinase by endothelin and noradrenaline in rat mesenteric arteries. Both vasoconstrictors activated p38alpha and/or p38beta but not p38gamma or p38delta, leading to increased HSP kinase activity. p38MAPK activation by noradrenaline was maximum between 2 and 10 minutes and was wholly dependent on calcium influx but insensitive to the tyrosine kinase inhibitor herbimycin A. In contrast, endothelin induced a biphasic response, with activation at 2 and 10 minutes. The early activity was wholly dependent on calcium influx and inhibited by herbimycin A. The later activity was only 50% calcium dependent, was insensitive to herbimycin A, but was 50% inhibited by genistein, a nonselective tyrosine kinase inhibitor. With both agonists, p38MAPK activity returned to basal by 30 minutes. SB203580, a p38MAPK inhibitor, blocked agonist-induced HSP kinase activity, and herbimycin A inhibited activation by endothelin but not by noradrenaline. In addition, SB203580 inhibited noradrenaline-induced contraction but had little effect on contraction to endothelin. These data show that vasoconstrictors use different upstream activators of p38MAPK in vascular tissue and that the p38MAPK pathway is selectively implicated in the contractile response to noradrenaline in small arteries.

Animals↗
PubMed1998

Lipid second messengers derived from glycerolipids and sphingolipids, and their role in smooth muscle function.

The processes that link activation of an external receptor to the internal mechanisms that elicit a physiological response have been the subject of extensive investigation. It has been established that rather than just being an inert barrier to protect the cell from environmental damage, there are populations of phospholipids located within the plasma membrane that act as a reservoir for signalling molecules and when a receptor binds its appropriate activating ligand a chain of events is initiated which leads to the breakdown of these lipids and the release of second messengers. Such processes are rapid enough for physiological responses to be effected. The purpose of this review is to examine the profile of lipid second messengers derived from glycerophospholipids and sphingolipids. In the former class are included phosphoinositide and phosphatidylcholine and the latter includes sphingomyelin. Hydrolysis of such parent compounds is mediated by phospholipases and the profile of metabolites appears to be agonist specific and modulated by a number of mechanisms including heterotrimeric G-protein subunits, small G-proteins, alterations in intracellular calcium concentration, protein kinase C and tyrosine kinases. The recent interest in sphingolipids, particularly in vascular smooth muscle cells, has been provoked by the observation that ceramide and sphingoid base formation is observed in response to vasoconstrictor hormones.

Animals↗
PubMed1997

Involvement of tyrosine phosphorylation in endothelin-1-induced calcium-sensitization in rat small mesenteric arteries.

1. We have studied the effect of endothelin-1 stimulation on protein tyrosine phosphorylation levels in intact small mesenteric arteries of the rat and investigated the effects of tyrosine kinase inhibition on the contractile response to this agonist. 2. Endothelin-1 stimulated a rapid (20 s), sustained (up to 20 min) and concentration-dependent (1-100 nM) increase in protein tyrosine phosphorylation levels which coincided temporally with the contractile response in intact and alpha-toxin permeabilized small artery preparations. Tyrosine phosphorylation was increased in four main clusters of proteins of apparent molecular mass 28-33, 56-61, 75-85 and 105-115 kDa. Endothelin-1-induced protein tyrosine phosphorylation was independent of extracellular calcium, antagonized by the tyrosine kinase inhibitor tyrphostin A23 but not by the inactive tyrphostin A1. 3. In intact small arteries tyrphostin A23 inhibited the force developed to endothelin-1 at all concentrations studied; at higher concentrations (10 and 100 nM) the profile of contraction was altered from a sustained to a transient response. Tyrphostin A1 inhibited the contractile response to endothelin-1 at all concentrations except 100 nM; the profile of the response was not altered. Neither tyrphostin affected the transient phasic contraction induced by endothelin-1 (100 nM) in the absence of extracellular calcium. 4. In rat alpha-toxin permeabilized mesenteric arteries endothelin-1 caused a concentration-dependent increase in force in the presence of 10 microM GTP and low (pCa 6.7) constant calcium, demonstrating increased sensitivity of the contractile apparatus to calcium. Tyrphostin A23 inhibited this response by approximately 50%, tyrphostin A1 did not affect endothelin-1-induced calcium sensitization of force. 5. We conclude that increased tyrosine phosphorylation is important in the contractile response induced by endothelin-1 in intact small mesenteric arteries. Furthermore our data implicate activation of this signalling pathway in the tonic phase of contraction possibly through modulation of the sensitivity of the contractile apparatus to calcium.

Animals↗
PubMed1997

Calcium sensitivity and agonist-induced calcium sensitization in small arteries of young and adult spontaneously hypertensive rats.

The sensitivity of the myofilaments to Ca2+ is increased during agonist-induced contraction of vascular smooth muscle. Given the important contribution of vascular tone to the elevation of peripheral resistance observed in genetic hypertension, we have investigated whether alterations in myofilament Ca2+ sensitivity occur in small arteries from spontaneously hypertensive rats (SHR) and age-matched Wistar-Kyoto (WKY) controls during the developmental and established phases of hypertension. Segments of mesenteric, renal, and femoral artery with an average lumen diameter <300 microm from 5- or 20-week-old rats were mounted in a wire myograph. Morphological measurements were made and the vessels permeabilized with Staphylococcus aureus alpha-toxin. Dose-response curves to increasing concentrations of Ca2+ were obtained and the ability of 100 nmol/L endothelin-1 (ET-1) or 10 micromol/L norepinephrine (NE) in the presence of 10 micromol/L GTP to enhance tension in response to low Ca2+ (pCa6.7) was determined. Systolic, diastolic, and mean blood pressures were higher in SHR than in WKY at 5 and 20 weeks. The media thickness:lumen diameter ratio was increased in mesenteric and femoral arteries from SHR compared with WKY at 5 and 20 weeks. There was no difference in media thickness:lumen diameter ratio in renal arteries or between 5- and 20-week animals in any vascular bed. The pCa curves were not different in mesenteric, renal, or femoral arteries from hypertensive compared with normotensive rats or between age groups, except in femoral arteries at 20 weeks, which exhibited a greater sensitivity to Ca2+ in SHR. Tension developed in response to maximal Ca2+ (pCa5.0) was greater in permeabilized mesenteric arteries from SHR compared with WKY at 20 weeks of age only; media stress was again similar in both strains but increased in older animals compared with younger animals in mesenteric arteries from WKY. The submaximal contraction induced by pCa6.7 was greater in femoral and renal than mesenteric arteries. GTP (10 micromol/L) augmented the tension developed to pCa6.7 in mesenteric arteries at 5 and 20 weeks and in renal arteries at 20 weeks. Addition of 100 nmol/L ET-1 or 10 micromol/L NE in the continued presence of GTP markedly increased tension in mesenteric arteries at 5 and 20 weeks. In renal arteries, 10 micromol/L NE enhanced Ca2+ sensitivity in the presence of GTP in SHR at 5 and 20 weeks and WKY at 5 weeks. In femoral arteries, there was a tendency for ET-1 and NE to increase Ca2+ sensitivity, but this increase was significant in WKY at 20 weeks (ET-1) and SHR at 5 weeks (NE) only. We have demonstrated that the sensitivity of the myofilaments to Ca2+ and ET-1- or NE-induced Ca2+ sensitization is not different in permeabilized small mesenteric, renal, or femoral arteries from SHR compared with WKY controls. Only in SHR mesenteric arteries at 20 weeks of age was there evidence of increased active tension in response to maximal Ca2+, despite structural differences, consistent with increased muscle mass in femoral arteries from SHR. We conclude that it is unlikely that a ubiquitous abnormality of the sensitivity of the contractile apparatus to Ca2+ or agonist-induced Ca2+ sensitization in vascular smooth muscle underlies the elevated total peripheral resistance associated with hypertension.

Aging↗
PubMed1996

Increase by lysophosphatidylcholines of smooth muscle Ca2+ sensitivity in alpha-toxin-permeabilized small mesenteric artery from the rat.

1. Pharmacological characterization of different lysophosphatidylcholines was performed based on their effect on the Ca2+ sensitivity of contraction in alpha-toxin-permeabilized rat mesenteric arteries. Furthermore, the effect of noradrenaline on [3H]-myristate-labelled lysophosphatidylcholine levels was assessed, to investigate whether lysophosphatidylcholines could be second messengers. 2. Palmitoyl or myristoyl L-alpha-lysophosphatidylcholine increased the sensitivity to Ca2+, whereas lysophosphatidylcholines containing other fatty acids had less or no effect. 3. L-alpha-phosphatidylcholine, L-alpha-glycerophosphorylcholine, palmitic acid, myristic acid and choline, potential metabolites of lysophosphatidylcholines, did not affect contractions. 4. Noradrenaline (GTP was required) and GTP gamma S increased the sensitivity to Ca2+, and GDP-beta-S inhibited the effect of noradrenaline. Lysophosphatidylcholines, however, had no requirement for GTP and caused sensitization in the presence of GDP-beta-S. 5. Calphostin C, a relatively specific protein kinase C inhibitor, did not affect contraction induced by Ca2+, but abolished the sensitizing effect of lysophosphatidylcholine. 6. Noradrenaline caused no measurable changes in the levels of [3H]-myristate-labelled phosphatidylcholine and lysophosphatidylcholine at 30 s and 5 min stimulation. 7. These results suggest that lysophosphatidylcholines can increase Ca2+ sensitivity through a G-protein-independent, but a protein kinase C-dependent mechanism. However, the role for lysophosphatidylcholines as messengers causing Ca2+ sensitization during stimulation with noradrenaline remains uncertain because no increase in [3H]-myristate labelled lysophosphatidylcholine could be measured during noradrenaline stimulation.

Animals↗
PubMed1996

Identification of protein kinase C isoforms in rat mesenteric small arteries and their possible role in agonist-induced contraction.

We have identified immunologically the protein kinase C (PKC) isoforms present in rat mesenteric small arteries, defined their distribution between particulate and soluble fractions, and studied their involvement in phorbol ester-induced contraction. Our analysis revealed the presence of the CA(2+)-dependent PKCs (alpha and gamma), Ca(2+)-independent PKCs (delta and epsilon), and the atypical isoform (zeta). PKCbeta could not be detected, whereas PKCgamma is likely to be of neural origin. All isoforms exhibited different distributions. PKCalpha, PKCepsilon, and PKCzeta were found in both particulate and soluble fractions. In contrast, PKCdelta was mainly in the particulate fraction, and PKCgamma was in the soluble fraction. Phorbol esters, which activate PKC and cause smooth muscle contraction, downregulated only the alpha and delta isoforms. This was associated with a parallel loss of contractile response to phorbol ester. The force developed to submaximal concentrations of noradrenaline was decreased after phorbol dibutyrate pretreatment, although the sensitivity and maximal response were unchanged. Phorbol ester pretreatment did not affect the contractile response to vasopressin. The sensitivity to non-receptor-mediated contraction, caused by k+ in the presence of prazosin, was slightly reduced by 4 alpha- and 4 beta-phorbol ester pretreatment. Maximal tension in response to this agonist was not affected. We conclude that PKCalpha and/or PKCdelta is necessary for phorbol ester-mediated contraction but is not essential for noradrenaline-, vasopressin-, or k(+)-induced contraction, demonstrating differences in the mechanisms involved in the contractile response between these agents.

Animals↗
PubMed1995

Phospholipase D-induced phosphatidate production in intact small arteries during noradrenaline stimulation: involvement of both G-protein and tyrosine-phosphorylation-linked pathways.

To investigate membrane lipid metabolism during smooth-muscle activation, the role of phospholipase D (PLD) in the production of phosphatidate (PA) was studied in rat small arteries stimulated with noradrenaline. Incubation with [3H]myristate preferentially labelled phosphatidylcholine (PtdCho), and in the presence of 0.5% ethanol [3H]phosphatidylethanol ([3H]PEt) was formed, demonstrating PLD activity. Noradrenaline (NA) stimulation resulted in an increase in PtdCho derived [3H]PA and [3H]PEt formation, indicating PLD activation. Stimulation of [14C]choline release confirmed PLD-mediated hydrolysis of PtdCho. Propranolol, an inhibitor of PA phosphohydrolase, increased [3H]PA levels in non-stimulated tissue and decreased the rate of degradation of both [3H]PA and [3H]PEt, implying that this is an active route for PA metabolism in small arteries. However, [3H]diacylglycerol levels were not increased during NA stimulation. Fluoroaluminate increased [3H]PEt formation and [14C]choline release, whereas high K+ in the presence of alpha 1-adrenoceptor blockade did not. Pervanadate increased phosphotyrosine levels in small arteries, and markedly stimulated [3H]PEt formation and [14C]choline release. The combination of pervanadate and NA stimulation resulted in a dramatic increase in [3H]PEt formation, which was greater than the sum of the individual responses to the two agonists. Pervanadate and fluoroaluminate in combination appeared to give an additive response, whereas high K+ did not alter the pervanadate-induced formation of [3H]PEt. Phosphotyrosine levels were increased by NA in the presence of tyrosine phosphatase inhibitors. This effect was blocked by genistein, a tyrosine kinase inhibitor. These data demonstrate that in NA-stimulated small arteries PLD-induced PtdCho hydrolysis contributes to accumulation of PA, but not of diacylglycerol. Furthermore, regulation of PLD activity appears to require G-protein and tyrosine-phosphorylation-linked pathways.

Animals↗
PubMed1995

Ultraviolet radiation stimulates a biphasic pattern of 1,2-diacylglycerol formation in cultured human melanocytes and keratinocytes by activation of phospholipases C and D.

Ultraviolet radiation (UVR) induces melanin synthesis by human epidermal melanocytes, and phospholipid-derived 1,2-diacylglycerols (DAGs) have been implicated in mediating this response. In previous experiments, addition of the synthetic DAG 1-oleoyl-2-acetylglycerol to cultured pigment cells stimulated melanogenesis. The purpose of the present study was to analyse the effects of UVR on the endogenous generation of DAGs. It was found that in a number of cultured cell types, including human melanocytes and B16 mouse melanoma cells, but also human keratinocytes and Swiss 3T3 fibroblasts, exposure to a single dose of UVR stimulated a biphasic increase in endogenous DAG formation. An early transient rise, over seconds, was followed by a more sustained delayed rise over minutes. The early rise in DAG levels was accompanied by a transient rise in inositol trisphosphate formation, indicating activation of phosphatidylinositol-specific phospholipase C. The delayed rise was accompanied by activation of phospholipase D. This endogenous DAG formation by pigment cells is further evidence for the involvement of DAGs in UVR-induced epidermal melanin synthesis. Since DAG formation is also seen in other cells types, it is possible that DAGs may be involved in an array of UVR-induced responses.

Cells, Cultured↗
PubMed1994

Membrane-associated diacylglycerol kinase activity is increased by noradrenaline, but not by angiotensin II, in arterial smooth muscle.

In rat small arteries, noradrenaline stimulates the sustained production of arachidonoyl-phosphatidic acid, whereas there is only a slight and transient increase with angiotensin II [Ohanian, Ollerenshaw, Collins and Heagerty (1990) J. Biol. Chem. 265, 8921-8928]. Diacylglycerol kinase (DGK) is the enzyme responsible for generating phosphatidic acid from 1,2-diacylglycerol (DAG). To investigate whether agonists influence DGK activity, we have studied this enzyme in both particulate and soluble fractions prepared from rat small arteries. Soluble DGK activity was inhibited by octyl glucoside. Therefore a deoxycholate assay was used for this fraction, whereas an octyl glucoside mixed-micelle assay was used to examine particulate fractions. Particulate DGK selectively phosphorylated long-chain DAG at a rate 2.5-3-fold higher than that for the synthetic substrate dioctanoylglycerol. In contrast, the substrate preference of the soluble isoenzyme(s) was: dioctanoylglycerol > arachidonoyl-DAG= dioleoylglycerol. Stimulation of intact arteries with noradrenaline (15 microM) increased membrane-associated DGK activity 3-fold, transiently. Angiotensin II (100 nM) stimulation did not alter the DGK activity of this fraction. The activity of the soluble DGK was increased by both agonists, but only transiently. These results demonstrate that rat small arteries contain a membrane-associated DGK which metabolizes arachidonoyl-containing substrate. Also, the activity of this enzyme is regulated differentially by vasoconstrictor hormones. It is concluded that modulation of DGK activity may represent one point at which agonists using the same signal-transduction pathway may tailor the cellular response.

Angiotensin II↗
PubMed1993

Regulation of diacylglycerol metabolism by vasoconstrictor hormones in intact small arteries.

The initiation of receptor-mediated small artery contraction is dependent on inositol 1,4,5-trisphosphate-stimulated release of stored calcium. The role of the other product of inositol lipid hydrolysis, 1,2-diacylglycerol, in maintaining contraction remains controversial. Therefore, we have determined the contractile response of rat subcutaneous small arteries (< 300 microns i.d.), when mounted as ring preparations in a myograph, to noradrenaline, angiotensin II, KCl-induced membrane depolarization, and a cell-permeable diglyceride, dioctanoylglycerol. In parallel experiments, the conversion of this diglyceride to dioctanoylphosphatidate was studied in 32P-labeled vessels. Dioctanoylglycerol produced a slow-onset sustained contraction that was dependent on extracellular calcium. This was accompanied by the generation of the lipid dioctanoylphosphatidate. Noradrenaline and KCl induced rapid-onset sustained contractions and increased the production of dioctanoylphosphatidate (75% and 91%, respectively). In addition, dioctanoylglycerol levels were reduced (41%) after noradrenaline stimulation, suggesting activation of diacylglycerol kinase. In contrast, the contractile response to angiotensin II was transient, and this agonist did not significantly affect the conversion of dioctanoylglycerol to phosphatidate. Noradrenaline markedly increased (fourfold) the formation of endogenous phosphatidate, whereas endogenous 1,2-diacylglycerol was increased (47%) with angiotensin II. These results demonstrate that phosphatidate formation is regulated by vasoconstrictor hormones during receptor-mediated contraction, independent of diglyceride mass. Modulation of the levels of lipid second messengers downstream from phospholipid hydrolysis may represent a mechanism by which agonists that act through the same signaling system produce different contractile responses.

Angiotensin II↗
PubMed1992

The phosphoinositide signaling system and hypertension.

The phosphoinositide signaling system is common to many vasoconstrictor agents and as such is influential in the regulation of blood pressure. Recently, there have been major advances in our understanding of these lipids and their metabolism. Characterization of the phospholipase C isozymes and protein kinase C isozymes involved in transmembrane signaling has progressed rapidly. The role of diacylglycerol kinase as a regulator of protein kinase C activity has been established, and phosphatidic acid has been recognized as a cellular messenger. Studies in the spontaneously hypertensive rat have shown abnormalities of phospholipase C that could result in enhanced activity and explain changes in sensitivity reported in rats with this disease. During agonist activation of inositol lipid hydrolysis, levels of inositol 1,4,5-trisphosphate and 1,2-diacylglycerol are elevated in spontaneously hypertensive rats compared with Wistar-Kyoto control rats. These changes are observed early, prior to blood pressure stabilization, and may be downregulated once hypertension is established. In addition, there is evidence for reduced diacylglycerol kinase activity and enhanced protein kinase C activity in the spontaneously hypertensive rat. These data provide evidence for hyperresponsiveness of the phosphoinositide signaling system in the developmental stages of hypertension. However, confirmatory experiments in nongenetic animal models of hypertension and in human tissues are needed to establish that this is not just a phenotypic phenomenon of the spontaneously hypertensive rat.

Animals↗
PubMed1990

Agonist-induced production of 1,2-diacylglycerol and phosphatidic acid in intact resistance arteries. Evidence that accumulation of diacylglycerol is not a prerequisite for contraction.

The production of total amounts of 1,2-diacylglycerol as well as those specifically derived from inositol lipid hydrolysis was studied in intact rat resistance arteries stimulated with either noradrenaline, vasopressin, or angiotensin II at 20 s when the onset of contraction would be nearing its maximum, and at 5 min during the sustained phase of contraction. Total amounts of 1,2-diacylglycerol were not altered by any agonist at 20 s, or at 5 min. However, arachidonate-containing species of 1,2-diacylglycerol were differentially influenced being increased at 5 min by noradrenaline, and decreased at 20 s and 5 min by vasopressin. Only angiotensin II produced substantial increases in this class of 1,2-diacylglycerol at both time points. In order to investigate the fate of this second messenger total and inositol lipid derived phosphatidic acids were then measured at both 20 s and 5 min. Noradrenaline induced a rise in both total and arachidonate-containing phosphatidic acid at both times as did vasopressin. Only small increases were induced by angiotensin II at 20 s. These data demonstrate that the accumulation of 1,2-diacylglycerol generated from inositol lipid breakdown is only observed with activation by angiotensin II. Other agonists produced phosphatidic acids with time and the rate of generation of these lipids is agonist-specific. Thus phosphatidic acid may play a more prominent role during the sustained phase of contraction than previously anticipated.

Angiotensin II↗
PubMed1990

Atrial natriuretic factor-induced systemic vasoconstriction in conscious dogs, rats, and monkeys.

This study addresses the hypothesis that atrial natriuretic factor (ANF) is a primary vasodilator, which reduces arterial pressure directly and increases total peripheral resistance secondarily by reflex mechanisms. The effects of 30-minute infusions of ANF (0.3 micrograms/kg/min i.v.) were examined in conscious dogs, rats, and monkeys before and after ganglionic blockade with hexamethonium. In seven intact, conscious dogs, ANF reduced mean arterial pressure by 7 +/- 1% and cardiac output by 19 +/- 3% and increased total peripheral resistance by 15 +/- 3%. After ganglionic blockade, ANF reduced mean arterial pressure by 7 +/- 2% but still increased total peripheral resistance by 15 +/- 3%. Similar results were observed in four dogs with total cardiac denervation and in six dogs with arterial baroreceptor denervation. Furthermore, in two dogs, combined ganglionic and alpha 1-adrenoceptor blockades failed to alter the rise in total peripheral resistance observed with ANF. In six intact, conscious rats, ANF reduced mean arterial pressure by 8 +/- 2% and cardiac output by 27 +/- 2% and increased total peripheral resistance by 27 +/- 5%. After ganglionic blockade, ANF still increased total peripheral resistance by 13 +/- 3%. In six intact, conscious monkeys, ANF reduced mean arterial pressure by 14 +/- 2% and cardiac output by 26 +/- 3% and increased total peripheral resistance by 17 +/- 3%. However, after ganglionic blockade. ANF decreased total peripheral resistance by 11 +/- 2%. These data provide evidence for a fundamental species difference in the vascular actions of ANF. In conscious dogs, ANF elicits "direct" vasoconstriction, which increases total peripheral resistance, even in the presence of denervation of reflexes or autonomic blockade. In conscious rats, ANF elicits both direct and reflexly mediated vasoconstriction. In conscious monkeys, although a component of direct vasoconstriction may also be present, the most prominent component appears to be reflexly mediated, since it was abolished by ganglionic blockade.

Adrenergic alpha-Antagonists↗