Biomedical subjects

Lars Edvinsson

Publications and source records attributed to Lars Edvinsson.

At least 73 records · Page 4Linked to original sources

PubMed2003

Subarachnoid hemorrhage enhances endothelin receptor expression and function in rat cerebral arteries.

OBJECTIVE: Inspired by organ culture-induced changes in the vascular endothelin (ET) receptor population, we investigated whether such changes occur in cerebral arteries in a rat subarachnoid hemorrhage (SAH) model. METHODS: SAH was induced with injection of 250 microl of blood into the prechiasmatic cistern. After 2 days, the middle cerebral artery, basilar artery, and posterior communicating artery were harvested. Pharmacological studies were performed in vitro, and levels of messenger ribonucleic acid (mRNA) were quantified in real-time reverse transcriptase-polymerase chain reaction assays. RESULTS: In the middle cerebral artery and basilar artery from rats with induced SAH, enhanced biphasic responses to ET-1 were observed. The -log(50% effective concentration) value for the high-affinity phase was approximately 12, compared with approximately 8.5 for sham-operated animals. At a concentration of ET-1 of 10(-9) mmol/L (approximately equal to the physiological concentration of ET-1 in the plasma), submaximal contractions of 50 to 75% of the contraction obtained through stimulation with 60 mmol/L K(+) were now observed. Quantitative mRNA studies with the same arteries demonstrated significant increases in the number of copies of ET(B) receptor mRNA but not ET(A) receptor mRNA. Evidence of functional ET(B) receptors was provided in antagonist studies. The posterior communicating artery did not exhibit significant changes. CONCLUSION: The altered receptor profile observed may represent the final stage in the series of events leading from SAH to actual spasm of the artery. The pharmacological data for the ET(B) receptor suggest complex interactions between normally present ET(A) receptors and up-regulated ET(B) receptors.

Animals↗

PubMed2002

Analysis of the time course for organ culture-induced endothelin ET B receptor upregulation in rat mesenteric arteries.

In contrast to the constitutively expressed endothelin ET(A) receptor, the distribution of endothelin ET(B) receptors is more variable. The aim of the present study was to investigate the kinetics of organ culture-induced upregulation of contractile endothelin ET(B) receptors in rat mesenteric arteries at both mRNA and functional levels. Assessment of mRNA expression revealed low levels of endothelin ET(B) receptor mRNA relative to endothelin ET(A) receptor mRNA after 3 h of culture, which gradually increased to reach a plateau level after 24 h. Correspondingly, vessels cultured for 3 h showed a negligible contractile response the selective endothelin ET(B) receptor agonist sarafotoxin 6c. Subsequently, the contractile response to sarafotoxin 6c was successively increased during organ culture until 24 h and, thereafter, a further increase in potency was seen after 48 h. These results demonstrate a rapid induction of transcription within less than 7 h followed by an increase in the response to receptor stimulation.

Animals↗

PubMed2002

Protein kinase C inhibitors decrease endothelin ET(B) receptor mRNA expression and contraction during organ culture of rat mesenteric artery.

The effect of protein kinase C (PKC) inhibitors on the induction of endothelin ET(B) receptors during organ culture was examined in isolated segments of the rat mesenteric artery. After 24 h of organ culture, the endothelin ET(B) receptor agonist sarafotoxin 6c (S6c) induced a strong contraction compared to fresh segments. The contractile response after 24-h organ culture to S6c was studied in presence (30-min preincubation) or absence, after 24-h treatment, of the PKC inhibitors staurosporine, K252a and Ro31-7549. Exposure to staurosporine or K252a in presence and after 24-h treatment reduced the S6c contraction. In contrast, presence of 2-1[1-3(aminopropyl)indol-3-yl]-3(1-methyl-1H-indol-3-yl)maleimide (Ro31-7549), did not affect the S6c-induced contraction, whereas 24-h treatment abolished the increase of contraction. The PKA inhibitor N-(2-[bromocinnamylamino]-ethyl)-5-isoquinolinesulfonamide (H89) did not affect the S6c responses. The mRNA expressions of endothelin ET(B) receptors (analysed with real-time PCR) were abolished after 24-h treatment with the PKC inhibitors. These results suggest that PKC is involved in the endothelin ET(B) receptor upregulation following organ culture.

Animals↗

PubMed2002

Capsaicin receptor immunoreactivity in the human trigeminal ganglion.

The cloned capsaicin receptor, also known as vanilloid receptor subtype 1 (VR1) receptor, has been demonstrated to be an integral membrane protein with homology to a family of putative store-operated calcium channels. The VR1 receptor is activated not only by capsaicin but also by noxious heat and protons, and therefore it is suggested as a molecular integrator of chemical and physical stimuli that elicit pain. In the present study, indirect immunofluorescence detected a small number of neurons that are VR1 receptor immunoreactive (ir) (171 versus 1038 or 16% of all neuronal cell bodies) in the human trigeminal ganglion (TG). In addition, RT-PCR confirmed the presence of VR1 mRNA in the human TG. It has been hypothesized that TG neuronal cell bodies are the source of capsaicin-stimulated release of calcitonin gene-related peptide (CGRP), and hence co-localization experiments were performed. Around 10% of the VR1 receptor-ir is expressed on neurons that contain CGRP-ir (ten among 74) in the human TG, indicating that capsaicin may act through the VR1 receptor to modulate the release of CGRP and in turn to modulate pain. We observed that 8% of the VR1 receptor-ir neuronal cell bodies contain substance P-ir and 5% nitric oxide synthase. Capsaicin can release nitric oxide, CGRP and substance P from sensory nerves and contribute to central sensitization.

Adult↗

PubMed2002

Reduction of bFGF-induced smooth muscle cell proliferation and endothelin receptor mRNA expression by mevastatin and atorvastatin.

The anti-atherosclerosis mechanisms of 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (statins) occur via both cholesterol-dependent and cholesterol-independent mechanisms. The present study used aortic and cerebral vascular smooth muscle cells (SMC) from rat to investigate whether atorvastatin and mevastatin affect basic fibroblast growth factor (bFGF)-induced SMC proliferation and the mRNA expression of endothelin A (ET(A)) and endothelin B (ET(B)) receptors. Cell proliferation was assessed by MTT and real-time PCR was used to quantify ET(A) and ET(B) receptor mRNA. bFGF-induced concentration and time dependent SMC proliferation and up-regulation of the mRNA expression of ET(A) and ET(B) receptors. The 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors inhibited bFGF-induced proliferation of SMC (P<0.01). In aortic SMC atorvastatin and mevastatin significantly inhibited bFGF-induced mRNA expression of endothelin ET(A) and ET(B) receptors (P<0.05). Although in cerebral SMC the inhibitory effect of the statins was comparable in size with that seen in aortic SMC, only reached borderline significance (P=0.06) for ET(A) receptor mRNA but not for ET(B). The findings suggested a direct effect of statins on the vascular wall beyond their well-known lipid lowering effect in anti-atherosclerosis. Furthermore, the specific antagonists of ET(A) and ET(B) receptors (FR139317 and BQ788, respectively) significantly inhibited bFGF-induced SMC proliferation (P<0.001). The results suggested that endothelin receptors and the mevalonate pathway were involved in bFGF-induced SMC proliferation.

Animals↗

PubMed2002

Calcitonin gene-related peptide (CGRP) in cerebrovascular disease.

Cerebral blood vessels are innervated by sensory nerves that store several neurotransmitters among which calcitonin gene-related peptide (CGRP) is the most abundant. In primary headaches, there is a clear association between the head pain and the release of CGRP. In cluster headache there is an additional release of vasoactive intestinal peptide (VIP). In connection with administration of triptans, the headache subsides and the neuropeptide release normalises, in part via a presynaptic effect. In subarachnoid hemorrhage (SAH), CGRP is released to counterbalance the blood-induced vasospasm. In severe cases, the stored CGRP may be exhausted while infusion of CGRP may limit cerebral vasospasm. Thus, interactions with the trigeminovascular system at CGRP receptors may be a useful target for understanding of cerebrovascular disease and to design novel treatments.

Animals↗

PubMed2002

Organ culture: a new model for vascular endothelium dysfunction.

BACKGROUND: Endothelium dysfunction is believed to play a role in the development of cardiovascular disease. The aim of the present study was to evaluate the suitability of organ culture as a model for endothelium dysfunction. METHODS: The isometric tension was recorded in isolated segments of the rat mesenteric artery branch, before and after organ culture for 20 h. Vasodilatation was expressed as % of preconstriction with U46619. The acetylcholine (ACh) induced nitric oxide (NO) mediated dilatation was studied in the presence of 10 microM indomethacin, 50 nM charybdotoxin and 1 microM apamin. Endothelium-derived hyperpolarising factor (EDHF) was studied in the presence of 0.1 mM L-NOARG and indomethacin. Prostaglandins were studied in the presence of L-NOARG, charybdotoxin and apamin. RESULTS: The ACh-induced NO and prostaglandin-mediated dilatations decreased significantly during organ culture (NO: 84% in control and 36% in cultured; prostaglandins: 48% in control and 16% in cultured). Notably, the total ACh-dilatation was not changed. This might be explained by the finding that EDHF alone stimulated a full dilatation even after organ culture (83% in control and 80% in cultured). EDHF may thereby compensate for the loss in NO and prostaglandin-mediated dilatation. Dilatations induced by forskolin or sodium nitroprusside did not change after organ culture, indicating intact smooth muscle cell function. CONCLUSIONS: Organ culture induces a loss in NO and prostaglandin-mediated dilatation, which is compensated for by EDHF. This shift in mediator profile resembles that in endothelium dysfunction. Organ culture provides an easily accessible model where the molecular changes that take place, when endothelium dysfunction is developed, can be examined over time.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗

PubMed2002

Distribution of mRNA for VIP and PACAP receptors in human cerebral arteries and cranial ganglia.

The distribution of mRNA for pituitary adenylate cyclase-activating polypeptide (PACAP) type 1 (PAC1) and vasoactive intestinal polypeptide (VIP) types 1 and 2 (VPAC1 and VPAC2, respectively) receptors was examined by reverse transcriptase polymerase chain reaction (RT-PCR) in human cerebral arteries and in trigeminal, otic, sphenopalatine and superior cervical ganglia. RT-PCR products of the expected sizes were detected in the arteries, in both the presence and absence of endothelium. In the majority (80%) of the trigeminal, otic and superior cervical ganglia, mRNA for all three receptors were found (n = 5). However, in the sphenopalatine ganglion neither VPAC2 nor PAC1 was detected (n = 2). This finding indicates the occurrence of both prejunctional (ganglia) and postjunctional (vessels) location of VIP and PACAP receptors.

Adult↗

PubMed2002

Effect of the CGRP receptor antagonist BIBN4096BS in human cerebral, coronary and omental arteries and in SK-N-MC cells.

Several lines of evidence suggest that a calcitonin-gene related peptide (CGRP) receptor antagonist may serve as a novel abortive migraine treatment. Here we present data on a human cell line and isolated human vessels for such an antagonist, BIBN4096BS. On SK-N-MC membranes, radiolabelled CGRP was displaced by both CGRP-(8-37) and BIBN4096BS, yielding pK(i) values of 8.5 and 11.4, respectively. Functional studies with SK-N-MC cells demonstrated that CGRP-induced cAMP production was antagonised by both CGRP-(8-37) and BIBN4096BS with pA(2) values of 7.8 and 11.2, respectively. Isolated human cerebral, coronary, and omental arteries were studied with a sensitive myograph technique. CGRP induced a concentration-dependent relaxation that was antagonized by both CGRP-(8-37) and BIBN4096BS in a competitive manner. CGRP was a weaker agonist on coronary arteries as compared to intracranial arteries; however, BIBN4096BS was an equally effective antagonist. In human omental arteries, CGRP did not induce relaxation. BIBN4096 had a pA(2) value of 10.1 in cerebral and 10.4 in coronary arteries. The results of clinical trials with BIBN4096BS for acute migraine attacks are awaited with great interest.

Calcitonin Gene-Related Peptide↗

PubMed2002

D-erythro-N,N-dimethylsphingosine inhibits bFGF-induced proliferation of cerebral, aortic and coronary smooth muscle cells.

The role of sphingosine kinase (SphK) on basic fibroblast growth factor (bFGF)-induced proliferation of cerebral, aortic and coronary smooth muscle cells (SMC) was addressed using D-erythro-N,N-dimethylsphingosine (DMS), an inhibitor of SphK which blocks conversion of sphingosine to sphingosine-1-phosphate (S1P). DMS concentration-dependently reduced the bFGF-induced proliferation of rat cerebral and aortic, and human coronary SMC. This suggests that SphK is one of the key enzymes in the mitogenic response to bFGF in vascular SMC as supported by the finding that S1P stimulated proliferation of SMC. Fumonisin B1, a dihydroceramidesynthase inhibitor which blocks the conversion of dihydrosphingosine to seramide, did not affect SMC proliferation induced by bFGF. Staurosporine, an inhibitor of protein kinase C (PKC), inhibited proliferation of SMC induced by bFGF, and both bFGF- and S1P-induced proliferation of SMC was sensitive to pertussis toxin (PTX), an inhibitor of Gi-protein activity. The present study thus demonstrates that SphK, PKC and Gi-protein activities are required for bFGF-mitogenic signaling in SMC. The bFGF mitogenic effect in vascular SMC might at least in part act via the SphK pathway and a Gi-protein.

Analysis of Variance↗

PubMed2002

Neuropeptide Y Y1 and neuropeptide Y Y2 receptors in human cardiovascular tissues.

mRNA encoding the human NPY Y1 and NPY Y2 receptors were detected in cerebral, meningeal, and coronary arteries using reverse transcriptase-polymerase chain reaction (RT-PCR). In addition, the trigeminal and superior cervical ganglia were positive for both receptors. In some arteries and in SK-N-MC cells only mRNA encoding the NPY Y1 was detected. Besides the expected NPY Y1 PCR products, an additional 97 bp longer amplicon originating from an alternative splicing event was found in most tissues studied. Antibodies directed against the NPY Y1 receptor revealed immunostaining mainly in the smooth muscle layer of blood vessels whereas antibodies against the NPY Y2 receptor showed immunostaining in nerve cell bodies.

Alternative Splicing↗

PubMed2002

Does nociceptin play a role in pain disorders in man?

Nociceptin-immunoreactive cellbodies were detected in the human trigeminal ganglion, while no such fibers were identified in the temporal artery or in dermal tissue from the neck region. In four healthy subjects receiving nociceptin into the temporal muscle in an open labeled design no pain was detected. In 10 healthy subjects who received 200pmol of nociceptin into tender non-dominant trapezius muscles in a placebo-controlled, randomized, balanced, and double-blinded design local tenderness increased (P=0.025) while no pain was noted. Thus, the action of nociceptin should be searched for in the trigeminal ganglion and/or in the central nervous system (CNS).

Adult↗

PubMed2002

Comparison of the antagonistic effects of different angiotensin II receptor blockers in human coronary arteries.

BACKGROUND: Angiotensin II (Ang II) is a potent vasoconstrictor and a deleterious factor in cardiovascular pathophysiology. Ang II receptor blockers (ARBs) have recently been introduced into clinical practice for treatment of hypertension and congestive heart failure. AIMS: This study was undertaken to evaluate the inhibitory effects of ARBs on vasoconstriction in humans. METHODS: Vasomotor tone was analyzed in endothelium denuded, human coronary artery (HCA) segments. Ang II type 1 (AT(1)) and type 2 (AT(2)) receptor mRNA expression was examined by reverse transcriptase-polymerase chain reaction (RT-PCR). RESULTS: Ang II was a potent vasoconstrictor (pEC(50) = 7.7). At 1 nM of the AT(1) receptor antagonists, candesartan and valsartan, the maximum contraction was depressed to 57 and 50% of Ang II, respectively, indicating insurmountability. Although generally considered surmountable, the presence of 100 nM losartan elicited a depression of the Ang II response to 32%. Its active metabolite, EXP 3174 (1 nM), abolished the Ang II contraction. The AT(1) receptor antagonists had the following order of blocking effect; EXP 3174 > candesartan = valsartan > losartan. The AT(2) receptor antagonist, PD 123319 (100 nM) significantly attenuated the Ang II contraction (E(max) = 62% of Ang II). RT-PCR of HCA smooth muscle cells demonstrated expression of both AT(1) and AT(2) receptor mRNA. CONCLUSIONS: Ang II contraction in HCA is mediated mainly by AT(1) but also involves AT(2) receptors. The active metabolite of losartan, EXP 3174, is the most efficacious AT(1) receptor antagonist in HCA.

Adult↗

PubMed2002

Protein kinase mediated upregulation of endothelin A, endothelin B and 5-hydroxytryptamine 1B/1D receptors during organ culture in rat basilar artery.

1. Organ culture has been shown to upregulate both endothelin (ET) and 5-hydroxytryptamine 1B/1D (5-HT(1B/1D)) receptors in rat cerebral arteries. The purpose of the present study was to investigate the involvement of protein kinases, especially protein kinases C (PKC) and A (PKA) in this process. 2. The effect of inhibiting protein kinases during organ culture with staurosporine (unspecific protein kinase inhibitor), RO 31-7549 (specific inhibitor of classical PKC's) and H 89 (specific inhibitor of PKA) was examined using in vitro pharmacological examination of cultured vessel segments with ET-1 (unspecific ET(A) and ET(B) agonist), S6c (specific ET(B) agonist) and 5-CT (5-HT(1) agonist). Levels of mRNA coding for the ET(A), ET(B), 5-HT(1B) and 5-HT(1D) receptors were analysed using real-time RT-PCR. 3. Classical PKC's are critically involved in the appearance of the ET(B) receptor; co-culture with RO 31-7549 abolished the contractile response (6.9 +/- 1.8%) and reduced the ET(B) receptor mRNA by 44 +/- 4% as compared to the cultured control. Correlation between decreased ET(B) receptor mRNA and abolished contractile function indicates upstream involvement of PKC. 4. Inhibition of PKA generally had an enhancing effect on the induced changes giving rise to a 7-25% increase in E(max) in response to ET-1, S6c and 5-CT as compared to the cultured control. 5. Staurosporine inhibited the culture induced upregulation of the response of both the ET(A) and the 5-HT(1B/1D) receptors, but had no significant effect on the mRNA levels of these receptors. This lack of correlation indicates an additional downstream involvement of protein kinases.

Animals↗

PubMed2002

Immunohistochemical localization of calcitonin receptor-like receptor and receptor activity-modifying proteins in the human cerebral vasculature.

Calcitonin gene-related peptide and adrenomedullin belong to a structurally related neuropeptide family and are potent vasodilators expressed in the trigeminovascular system. The molecular identity of receptors for these proteins has only recently been elucidated. Central to functional binding of these neuropeptides is the G-protein-coupled receptor, the calcitonin receptor-like receptor (CRLR), whose cell surface expression and pharmacology is determined by coexpression of a receptor activity-modifying protein (RAMP). CRLR combined with RAMP binds calcitonin gene-related peptide with high affinity, whereas CRLR coexpression with RAMP2 or -3 confers high-affinity binding of adrenomedullin. The authors investigated the expression of these receptor components in human cerebral vasculature to further characterize neuropeptide receptor content and the potential functions of these receptors. Localization has been carried out using specific antisera raised against immunogenic peptide sequences that were subsequently applied using modern immunohistochemical techniques and confocal microscopy. The results are the first to show the presence of these receptor component proteins in human middle meningeal, middle cerebral, pial, and superficial temporal vessels, and confirm that both calcitonin gene-related peptide and adrenomedullin receptors may arise from the coassembly of RAMPs with CRLR in these vessel types. These novel data advance the understanding of the molecular function of the trigeminovascular system, its potential role in vascular headache disorders such as migraine, and may lead to possible ways in which future synthetic ligands may be applied to manage these disorders.

Autoradiography↗

PubMed2002

Human endothelin subtype A receptor enhancement during tissue culture via de novo transcription.

OBJECTIVE: Endothelin (ET) has, since its discovery, increasingly been considered a key player in the pathophysiological processes of cerebral vasospasm in the course of subarachnoid hemorrhage, although it remains unclear how ET is involved. We present data that indicate an inherent capacity of human cerebral arteries to change their sensitivity to ET. METHODS: Human cerebral arteries were obtained from patients undergoing intracranial tumor surgery. The vessels were divided into segments and subjected to organ culture for 48 hours. The vessels were then examined by using in vitro pharmacological methods and molecular biological techniques. RESULTS: After organ culture of the cerebral arteries, both the sensitivity to and potency of ET were enhanced (maximal response, 152 +/- 9%; -log (50% effective concentration), 10.3 +/- 0.3), in comparison with data for fresh cerebral arteries. Contractions were inhibited by both FR139317 (a specific ET(A) receptor antagonist) and bosentan (a mixed ET(A) and ET(B) receptor antagonist), in a manner indicating the sole presence of contractile ET(A) receptors. An inconsistent dilative response to the selective ET(B) receptor agonist sarafotoxin 6c was observed; the response was preserved in some segments and abolished in others, and potentiation of the precontraction was observed in yet other segments. No isolated contractile response to sarafotoxin 6c was observed, however. In reverse transcription-polymerase chain reaction assays, both ET(A) and ET(B) receptor messenger ribonucleic acid was detected. CONCLUSION: These results demonstrate that human cerebral arteries are capable of enhancing the function of ET(A) receptors.

Cerebral Arteries↗

PubMed2002

UDP acts as a growth factor for vascular smooth muscle cells by activation of P2Y(6) receptors.

Mitogenic effects of the extracellular nucleotides ATP and UTP are mediated by P2Y(1), P2Y(2), and P2Y(4) receptors. However, it has not been possible to examine the highly expressed UDP-sensitive P2Y(6) receptor because of the lack of stable, selective agonists. In rat aorta smooth muscle cells (vascular smooth muscle cells; VSMC), UDP and UTP stimulated (3)H-labeled thymidine incorporation with similar pEC(50) values (5.96 and 5.69). Addition of hexokinase did not reduce the mitogenic effect of UDP. In cells transfected with P2Y receptors the stable pyrimidine agonist uridine 5'-O-(2-thiodiphosphate) (UDPbetaS) was specific for P2Y(6) with no effect on P2Y(1), P2Y(2), or P2Y(4) receptors. UDPbetaS stimulated [(3)H]thymidine and [(3)H]leucine incorporation and increased cell number in VSMC. Flow cytometry demonstrated that UDP stimulated cell cycle progression to both the S and G(2) phases. The intracellular signal pathways were dependent on phospholipase C, possibly protein kinase C-delta, and a tyrosine kinase pathway but independent of G(i) proteins, eicosanoids, and protein kinase A. The half-life of P2Y(6) receptor mRNA was <1 h by competitive RT-PCR. The mitogen-activated protein kinase kinase inhibitor PD-098059 significantly suppressed, whereas ATP and interleukin-1beta upregulated, expression of P2Y(6) receptor mRNA. The results demonstrate that UDP stimulates mitogenesis through activation of P2Y(6) receptors and that the receptor is regulated by factors important in the development of vascular disease.

Animals↗

PubMed2002

Cerebral ischemia upregulates vascular endothelin ET(B) receptors in rat.

BACKGROUND AND PURPOSE: Elevated levels of endothelin-1 (ET-1) have been reported in cerebral ischemia. A role for ET may prove more important if the vascular receptors were changed. We addressed whether there is any change in ET receptor expression in cerebral ischemia. METHODS: The right middle cerebral artery (MCA) was occluded in male Wistar rats for 2 hours with the intraluminal filament method. The basilar artery and both MCAs were removed after 46 hours of recirculation. The contractile responses to ET-1, a combined ET(A) and ET(B) receptor agonist, and sarafotoxin 6c (S6c), a selective ET(B) receptor agonist, were examined in vitro, and ET receptor mRNA was quantified by real-time polymerase chain reaction. RESULTS: S6c, which had no contractile effect per se on fresh or sham-operated rat cerebral arteries, induced a marked contraction in the occluded MCA (E(max) [maximum contraction, calculated as percentage of the contractile capacity of 63.5 mmol/L K+]=68+/-68%; P<0.0001), while there was no difference in the responses to ET-1 after cerebral ischemia. Real-time polymerase chain reaction revealed a significant upregulation of both the ET(A) and ET(B) receptors (both P<0.05) in the occluded MCA compared with the nonoccluded MCA from the same rats. CONCLUSIONS: Focal cerebral ischemia in rat induces increased transcription of both ET(A) and ET(B) receptors, which results in the appearance of a contractile response to the ET(B) receptor agonist S6c. These results suggest a role for ET receptors in the pathogenesis of a vascular component after cerebral ischemia.

Animals↗