Sociotropic cognition moderates stress-induced cardiovascular responsiveness in college women.

This study examined the moderating effects of sociotropic cognition (SC), a nondefensive need for approval, on stress-induced cardiovascular responsiveness (CVR) in women. Sixty-seven college-age females had blood pressure (BP) and heart rate (HR) monitored during baseline, anticipation, story-telling (where participants were randomly assigned to a low or high threat condition), and recovery periods. SC showed a positive association with CVR only in the high interpersonal threat context during task and early stages of the recovery periods. SC was positively correlated with such variables as anxiety, ruminative style, dysphoria, and anger. This is the first report examining the moderating effects of SC on interpersonal stress-induced CVR prior to, during, and following a task, using an explicit manipulation of social evaluation. The data help define risk factors for CVR in women, which may aid in the understanding of how emotions and stress affect physical health and well-being.

Low-density lipoprotein stimulated peroxide production and endocytosis in cultured human endothelial cells: mechanisms of action.

The effects of arachidonic acid metabolism and NADPH oxidase inhibitor on the hydrogen peroxide (H2O2) generation and endocytotic activity of cultured human endothelial cells (EC) exposed to atherogenic low-density lipoprotein (LDL) levels have been investigated. EC were incubated with 240 mg/dl LDL cholesterol and cellular H2O2 production and endocytotic activity measured in the presence and absence of the arachidonic acid metabolism inhibitors, indomethacin, nordihydroguaiaretic acid, and SKF525A, and NADPH oxidase inhibitor, apocynin. All inhibitors, with the exception of indomethacin, markedly reduced high LDL-induced increases in EC H2O2 generation and endocytotic activity. EC exposed to exogenously applied arachidonic acid had cellular functional changes similar to those induced by high LDL concentrations. EC incubated with 1-25 uM arachidonic acid had increased H2O2 production and heightened endocytotic activity. Likewise, EC pre-loaded with [3H]arachidonic acid when exposed to increasing LDL levels (90-330 mg/dl cholesterol) had a dose-dependent rise in cytosolic [3H]arachidonic acid. The phospholipase A2 inhibitors, 4-bromophenacyl bromide and 7,7-dimethyleicosadienoic acid, markedly inhibited H2O2 production in EC exposed to 240 mg/dl LDL cholesterol. These findings suggest that arachidonic acid contributes mechanistically to high LDL-perturbed EC H2O2 generation and heightened endocytosis. Such cellular functional changes add to our understanding of endothelial perturbation, which has been hypothesized to be a major contributing factor in the pathogenesis of atherosclerosis.

Comparison of neurons in rat medulla oblongata with fos immunoreactivity evoked by seizures, chemoreceptor, or baroreceptor stimulation.

Neurons in rat medulla oblongata with Fos immunoreactivity as a marker of synaptic excitation evoked by pentylenetetrazole-induced seizures were compared with cell populations activated by the stimulation of chemoreceptor and baroreceptor afferent pathways. Chemoreceptors were stimulated by placing rats in a hypoxic gas mixture (7% oxygen) for 2 h. Baroreceptors were activated by phenylephrine-induced hypertension. Seizures and hypoxia induced Fos immunoreactivity in neurons with similar anatomical distributions in the nucleus tractus solitarius, dorsal motor nucleus of the vagus, and ventrolateral medulla. Hypertension was associated with Fos immunoreactivity in an overlapping anatomical distribution compared to seizures and hypoxia, but in a more restricted pattern. A similar proportion of catecholaminergic cells of medulla oblongata (cells immunoreactive for catecholamine synthetic enzymes, tyrosine hydroxylase or phenylethanolamine-N-methyltransferase) had Fos immunostaining after seizures and hypoxia (P > 0.05), while significantly fewer were activated by hypertension (P < 0.05). The majority of tyrosine hydroxylase-immunoreactive cells in caudal ventrolateral medulla were activated by both seizures and hypoxia (mean per cents, 79 and 67%, respectively). Since cell populations activated by seizures and hypoxia are indistinguishable, and a majority of tyrosine hydroxylase-reactive cells in caudal ventrolateral medulla are independently activated by each stimulus, it may be inferred that some impulses originating from seizures and chemoreceptor afferent pathways converge to a common set of neurons. These observations identify neurons in rat medulla oblongata which may mediate the impact of seizures on central processing of chemoreceptor afferent activity.

In vivo effects of angiotensin II on vascular smooth muscle contraction and blood pressure are mediated through a protein tyrosine-kinase-dependent mechanism.

The effects of in vivo angiotensin II (AII) treatment on protein tyrosine kinase (PTK) levels were examined. It was found that administration of AII to normotensive Wistar-Kyoto rats induced tyrosine phosphorylation in aorta in a time-dependent manner. There was a rapid increase in phosphotyrosine content as early as 1 min, with peak phosphorylation occurring between 5 and 10 min. The response was also dose dependent, with increases in phosphorylation levels from 1 to 1000 micrograms/kg AII. Tyrosine phosphorylation was blocked using the angiotensin type 1 receptor antagonist losartan (10 mg/kg), suggesting that the effects are receptor mediated. Tyrphostin-25 (100 microM), a selective inhibitor of PTKs, when given in vivo was also able to attenuate phosphorylation by AII, further suggesting a PTK-mediated event. To couple these biochemical changes with physiological events, we also examined the ability of AII to induce vasoconstriction and raise systolic blood pressure through a PTK-mediated mechanism. In addition to increasing phosphorylation levels, AII caused a rise in systolic pressure in vivo and induced contraction in vitro. Both of these responses could be attenuated by pretreatment with losartan or tyrphostin-25. This is the first demonstration of the effects of AII on tyrosine phosphorylation in vivo. The data suggest that AII may induce a pressor response at least in part through activation of PTKs and subsequent phosphorylation of smooth muscle contractile proteins or activation of other protein kinases.

Tyrosine kinase inhibition suppresses angiotensin contraction in hypertensive and normotensive small resistance arteries.

Protein tyrosine kinases (PTKs) may influence vascular resistance, which is controlled primarily at the level of small arteries and arterioles. This study evaluates these kinases in resistance arteries from spontaneously hypertensive (SHR) and normotensive (WKY) rats using the protein tyrosine kinase inhibitor, tyrphostin-25. Gracilis muscle arteries (90-160 um, internal diameter) were mounted on a resistance vessel myograph and contractile responses to phenylephrine (PE) and angiotensin II (AII) were measured in the presence and absence of tyrphostin-25 (0.02-20 uM). Tyrphostin-25 inhibited AII, but not PE contractions and was less potent in the hypertensive arteries. This demonstrates for the first time that PTKs contribute to the contractile activity of resistance arteries from both hypertensive and normotensive rats. Further, results confirm in small arteries that AII-induced contractions are PTK-dependent and that arteries from hypertensive rats are hyporesponsive to PTK inhibition.

Alterations in splenocyte protein kinase C (PKC) activity by 2,3,7,8-tetrachlorodibenzo-p-dioxin in vivo.

The effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on growth factor-coupled activation of nuclear protein kinase C (nPKC) and on the subcellular distribution of PKC activity in rat splenocytes were investigated. Seven days after a single injection of TCDD (50 micrograms/kg body weight), cytosolic and particulate PKC activity was significantly higher in splenocytes from TCDD-treated rats or pair-fed control rats compared to ad libitum-fed animals. In a separate experiment, purified splenocyte nuclei from TCDD-treated animals and controls were used to study activation of nPKC by growth factors and other trophic agents. Growth factor-stimulated nPKC activation was attenuated in splenic nuclei from TCDD-treated rats compared to vehicle-treated controls. Evidence presented here suggests that the cellular mechanism of TCDD toxicity leading to immunosuppression in rodents may be mediated in part by uncoupling of growth factor receptors linked to PKC activation at the level of the nucleus. However, changes in total splenocyte PKC activity appear to be correlated with hypophagia since cytosolic and particulate PKC levels were elevated in TCDD-treated rats and their pair-fed partners.

Seizure-induced c-fos expression in rat medulla oblongata is not dependent on associated elevation of blood pressure.

This study was performed to determine whether c-fos immunoreactivity (IR) induced in medulla oblongata by pentylenetetrazole seizures is a consequence of seizure-associated blood pressure elevation and activation of baroreceptor afferent pathways, or occurs independently of hypertension. Immunohistochemical study of sections of medulla oblongata revealed that seizures are followed by induction of c-fos IR in nucleus tractus solitarius (NTS), dorsal motor nucleus of the vagus (DMN 10), and ventrolateral medulla (VLM), while there is negligible c-fos IR after saline sham injections. Seizures were associated with blood pressure elevation peaking at 31 +/- 17% (+/- SD) above baseline. Experimental hypertension at a similar level induced by i.p. phenylephrine also resulted in induction of c-fos IR in NTS. When seizures were preceded by antihypertensive treatment with the alpha-adrenergic antagonist, phentolamine, peak blood pressure tended to remain near the baseline level and lower than sham-injected controls. Normotensive seizures were associated with c-fos IR in NTS, DMN 10, and VLM similar to the pattern following hypertensive seizures. Seizure-induced activation of c-fos IR occurred despite normal blood pressure, and thus can be attributed to a direct effect of the seizure, and not to an indirect effect mediated by hypertension.

Retinoic acid induces translocation of protein kinase C (PKC) and activation of nuclear PKC (nPKC) in rat splenocytes.

Retinoic acid (RA), a vitamin A metabolite, has marked effects on growth of normal and malignant cells; however, the exact mechanism of action remains unclear. The effect of two RA analogs, 13-cis-RA and all-trans-RA, on transmembrane signalling processes was investigated in rat splenocytes. Treatment of rat splenic cells with these retinoic acid analogs resulted in translocation of protein kinase C (PKC) from the cytosol to the membrane. Previous studies have described nuclear RA receptors (RARs and RXRs) for several species and the biologic activity of RA has been shown to be mediated by specific interaction with these nuclear receptors. Thus, activation of nuclear pool(s) of protein kinase C (nPKC) by RA analogs was also studied. Rat splenocyte nuclei pure by enzymatic and electron microscope criteria demonstrated a biphasic pattern of bell-shaped curves for both cis- and trans-RA with maximum statistically significant peak of phosphate incorporation into endogenous substrates at 10(-16) M cis-RA and 10(-16)-10(-17) M trans-RA. A monoclonal antibody to PKC and the PKC inhibitors, H-7, sphingosine, and staurosporine, blocked the RA-stimulated nuclear phosphorylation. The ability of RA to activate cell membrane PKC resulting in an increase in particulate PKC activity correlates well with the activation of nPKC since the particulate fraction would include nuclear enzyme systems. This ability of RA to activate nPKC and possibly affect the growth status of a cell may provide a missing link to our understanding of the cellular sites of action for this vitamin.

Cell-free thymic extract from hypertensive rats induces hypertension in normotensive rats.

This study examined the role of the thymus in hypertension. Seven to eight week old, normotensive Wistar-Kyoto rats (WKYs) (systolic blood pressure 138 +/- 7 mm Hg) received a bolus injection of (1) WKY thymic extract (control), (2) spontaneously hypertensive rat (SHR) thymic extract, (3) SHR liver extract or (4) normotensive Sprague-Dawley rat (SD) thymic extract. Blood pressures of WKYs receiving SHR thymic rose significantly (p < 0.01) over an eight week period (168 +/- 6 mm Hg), while WKYs injected with WKY thymic extract (143 +/- 10), SHR liver extract (144 +/- 5) or SD thymic extract (138 +/- 4) showed no change in blood pressure. Thymuses from WKYs injected with SHR extract were significantly (p < 0.01) smaller than thymuses from WKYs injected with WKY extract. Aorta from WKYs administered SHR extract were significantly (p < 0.01) hyperresponsive to contractile agents, suggesting that immune dysfunction may lead to vascular damage as seen in several hypertensive models. The results suggest that hypertension can be transferred via an endogenous thymic factor, possibly a viral pathogen.

The anti-tumor agent, taxol, attenuates contractile activity in rat aortic smooth muscle.

Using 16-20 week old female spontaneously hypertensive rats (SHRs), the effects of the antitumor agent, taxol, on vascular reactivity were examined. Taxol significantly inhibited contraction induced by phenylephrine, angiotensin II, phorbol 12,13-dibutyrate and increasing concentrations of calcium. The data suggest that taxol does not augment hypersensitivity of the vascular, but instead attenuates contractile activity and may have important implications with respect to treatment of women with both cancer and cardiovascular disease.

Tyrphostin attenuates platelet-derived growth factor-induced contraction in aortic smooth muscle through inhibition of protein tyrosine kinase(s).

The effect of protein tyrosine kinase (PTK) inhibition on spontaneously hypertensive rat vascular smooth muscle contraction was examined in this study. By using isolated aortic strips, it was found that platelet-derived growth factor (PDGF) (0.1-1 nM), an activator of PTKs, elicited contraction with an EC50 of 0.25 +/- 0.08 nM. Treatment with tyrphostin (0.02-200 microM), a selective inhibitor of PTKs, caused a significant rightward shift of the concentration-response curves (P < .05). The IC50 for tyrphostin in the spontaneously hypertensive rat was calculated to be 9.5 +/- 4.2 microM. Tyrphostin also inhibited contractile activity in normotensive control Wistar-Kyoto rat aorta with an IC50 of 0.24 +/- 0.09 microM. Tyrphostin inhibited PDGF-induced contraction over a range of calcium concentrations, suggesting that it may oppose contraction through inhibition of calcium influx via PDGF-induced receptor-operated channel. However, KCl-mediated voltage-operated calcium channels were largely unaffected by tyrphostin, because it was unable to relax aortae which had been partially depolarized. Tyrphostin also had no significant antagonistic effect on contraction induced by phenylephrine or phorbol-12,13-dibutyrate. For both of these agents, contraction is mediated through activation of protein kinase C, which further alludes to the specificity of tyrphostin for PTKs. Treatment with 1 nM PDGF caused a significant stimulation of particulate/membrane PTK activity in the aorta. Tyrphostin attenuated PDGF-induced PTK activity in a concentration-dependent manner. The data suggest that PTKs may play a role in vascular smooth muscle contraction and that specific inhibition of PTK activity results in vasorelaxation. Furthermore, the results suggest that vasoconstriction and vascular smooth muscle cell proliferation may share common biochemical signalling pathways.

Decreased sensitivity of aorta from hypertensive rats to vasorelaxation by tyrphostin.

The role of protein tyrosine kinases (PTKs) in vascular smooth muscle (VSM) contraction was examined in spontaneously hypertensive rats (SHRs). Aorta from SHRs was hyperresponsive to PTK-mediated contraction relative to normotensive Wistar-Kyoto rats (WKYs). Aorta from SHR was also hyporesponsive to vasorelaxation by tyrphostin, a selective inhibitor of PTKs. Further, we found alterations in PTK activity in aorta from SHRs. PDGF stimulated PTK activity to a greater extent in the SHR. Tyrphostin inhibited PDGF-induced PTK stimulation in both strains, however, activity returned to basal levels in the WKY only. The results suggest that PTKs may be involved in VSM contraction and in the development of hypertension.

Gamma-interferon corrects aberrant protein kinase C levels and immunosuppression in the spontaneously hypertensive rat.

The effects of gamma-interferon (gamma-IFN) on protein kinase C (PKC) levels and immunosuppression in the spontaneously hypertensive rat (SHR) were examined. First, an abnormal PKC distribution was found in spleen, thymus and aorta from SHRs relative to normotensive controls. Biweekly injections of rat recombinant gamma-IFN (1000 U/kg) restored basal or resting PKC levels to those found in normotensive Wistar-Kyoto (WKY) rats. We also examined the effects of in vivo gamma-IFN treatment on nuclear PKC (nPKC) activation in purified, isolated splenocyte nuclei. It was found that basal nPKC levels were higher in untreated SHRs than gamma-IFN SHRs or WKYs. Also, while nuclei from untreated SHRs were relatively unresponsive to various immunoreactive substances and PKC activators, gamma-IFN treatment significantly restored activity. Last, the proliferative response to mitogen challenge of isolated splenocytes from untreated SHRs, gamma-IFN-treated SHRs and WKYs was studied. Although gamma-IFN treatment did not restore the proliferative response to that of WKYs, the mitogen response was significantly enhanced by treatment with gamma-IFN. The data show that gamma-IFN acts to restore normal immune function and corrects aberrant PKC levels and adds to the growing body of knowledge suggesting a role for immune dysfunction in the etiology of hypertension.

Prolactin induces growth-related gene expression in rat aortic smooth muscle in vivo.

We examined the effects of in vivo administration of prolactin on growth-related gene expression in aorta. Optimal mRNA expression for both the proto-oncogene, c-myc and ornithine decarboxylase occurred at 22 mg/kg prolactin. For c-myc, this was seen as early as 15 min. Prolactin-induced ornithine decarboxylase mRNA expression began at 1 h. The results confirm work showing induction of these genes by prolactin in tissues where it is mitogenic and supports a role for prolactin in the trophic response of vascular smooth muscle.

Prolactin induces proliferation of vascular smooth muscle cells through a protein kinase C-dependent mechanism.

The effects of prolactin (PRL) on A10 (aortic smooth muscle) cell proliferation were examined by measuring both [3H]thymidine incorporation and increases in cell number. PRL induced a significant proliferative response from 10(-11) to 10(-7) M, with optimal activity at 10(-10) M. PRL also enhanced platelet-derived growth factor (PDGF)-induced proliferation. The possibility that PRL induces proliferation through a protein kinase C (PKC)-mediated mechanism was also examined. PRL caused activation of PKC from 10(-12) to 10(-8) M. Antiserum to PRL, a monoclonal antibody directed against the PRL receptor and the immunosuppressive agent cyclosporine A, were able to inhibit PRL-induced proliferation and activation of PKC. The PKC inhibitors, staurosporine, sphingosine, and 1-(-5-iso-quinoline-sulfonyl)-2-methylpiperazine (H-7) also antagonized both proliferation and PKC activation. These data strongly suggest that PRL-induced A10 cell proliferation is mediated through the PKC pathway and that this may play a role in vascular smooth muscle cell hyperplasia, characteristic of the pathogenesis of cardiovascular diseases such as hypertension and atherosclerosis.

Atrial peptides inhibit protein kinase C-mediated contraction in rat aorta.

The effects of atriopeptin III on phorbol ester-induced contraction were examined in aorta from Wistar-Kyoto (WKY) and spontaneously hypertensive rats (SHR). Precontracted SHR aorta was less responsive than WKY aorta to atriopeptin III-induced relaxation. Additionally, SHR aorta had significantly greater basal and phorbol ester-stimulated protein kinase C (PKC) activity than WKY aorta. The altered PKC response as well as hyporesponsiveness to atriopeptin III in SHR aorta may contribute to an altered vascular response and to the pathogenesis of hypertension.

Atrial natriuretic peptides inhibit protein kinase C activation in rat aortic smooth muscle.

The effects of synthetic atrial natriuretic peptide and a cyclic guanosine 3',5'-monophosphate analogue, 8-bromo cGMP, on protein kinase C activation in rat aortic smooth muscle from spontaneously hypertensive and control Wistar-Kyoto rats were examined. Both atrial natriuretic peptide and 8-bromo cGMP inhibited protein kinase C activation by phenylephrine, an alpha 1-adrenoceptor agonist. When phorbol 12,13-dibutyrate was used, a substance which bypasses phospholipase C activation and directly activates protein kinase C, only atrial natriuretic peptide was effective in inhibiting activation. Additionally, it was found that spontaneously hypertensive rat aorta showed significantly greater basal and stimulated levels of protein kinase C in comparison with Wistar-Kyoto rat aorta. The results suggest first, a defective protein kinase C system in hypertensive rats and that atrial natriuretic peptide attenuates protein kinase C activation through cGMP-dependent and cGMP-independent mechanisms. This inhibition of protein kinase C activity may then lead to vasorelaxation.

Prolactin and known modulators of rat splenocytes activate nuclear protein kinase C.

Prolactin (PRL) and other trophic factors rapidly activate a nuclear pool(s) of protein kinase C (nPKC) in purified splenocyte nuclei. The PRL also enhanced [2-3H]glycerol incorporation into nuclear mono- and triacylglycerol. An assay was devised which not only probed the ability of the hormone to activate protein kinase C (PKC) but also demonstrated the presence of nuclear substrates. Using this methodology, a biphasic concentration-response curve to PRL was observed. Heterologous species of PRL and various growth factors also activated nPKC. The PRL-induced nPKC stimulation was antagonized by various immunomodulators, G protein-coupling inhibitors, PKC inhibitors, a calmodulin inhibitor, and a peripheral benzodiazepine agonist and antagonist. A monoclonal antibody to PKC, anti-rat PRL antiserum and a monoclonal anti-rat PRL receptor antibody antagonized PRL-induced PKC-dependent nuclear phosphorylation, further implicating nPKC and a PRL receptor-mediated activation process. Nuclear PKC may be a major target for trophic regulation in response to both positive and negative growth signals.

Adenosine activation of a nuclear pool of protein kinase C in rat splenocytes.

The stimulatory effects of adenosine analogues on a nuclear pool of protein kinase C (PKC) were examined using isolated rat splenocyte nuclei. Nuclear receptors met pharmacological criteria of A1 adenosine receptors including a potency profile in which cyclopentyladenosine (CPA), a selective A1 agonist, was more potent than 2-phenylaminoadenosine (2PAA), a selective A2 agonist. The selective A1 receptor agonist N6-1-(phenyl-2R-propyl) adenosine (R-PIA) activated PKC whereas the S diastereomer did not. The adenosine-induced PKC response could be attenuated using a monoclonal antibody to PKC, an A1 receptor antagonist, three known PKC inhibitors and pertussis toxin (PTX). The results suggest that adenosine may exert immunomodulatory effects through the activation of nuclear PKC.