Role of Ca2+ channels in NE-induced increase in [Ca2+]iand tension in fetal and adult cerebral arteries

1999 ◽  
Vol 277 (1) ◽  
pp. R286-R294 ◽  
Author(s):  
Wen Long ◽  
Yu Zhao ◽  
Lubo Zhang ◽  
Lawrence D. Longo
Keyword(s):  
Plasma Membrane ◽  
Cerebral Arteries ◽  
Channel Blockers ◽  
Developmental Age ◽  
Intracellular Ca ◽  
Lanthanum Ion ◽  
Near Term ◽  
Dose Dependent Decrease ◽  
Nonpregnant Adult

In vascular smooth muscle, elevation of agonist-induced intracellular Ca2+ concentration ([Ca2+]i) occurs via both Ca2+ release from intracellular stores and Ca2+influx across the plasma membrane. In the cerebral vasculature of the fetus and adult the relative roles of these mechanisms have not been defined. To test the hypothesis that plasma membrane L-type and receptor-operated Ca2+ channels play a key role in NE-induced vasoconstriction via alterations in plasma membrane Ca2+ flux and that this may change with developmental age, we performed the following study. In main branch middle cerebral arteries (MCA) from near-term fetal (∼140 days) and nonpregnant adult sheep, we quantified NE-induced responses of vascular tension and [Ca2+]i(by use of fura 2) under standard conditions in response to several Ca2+ channel blockers and in response to zero extracellular Ca2+. In fetal and adult MCA, maximal NE-induced tensions (g) were 0.91 ± 0.12 ( n = 10) and 1.61 ± 0.13 ( n = 12), respectively. The pD2 values for NE-induced tension were both 6.0 ± 0.1, whereas the fetal and adult maximum responses (%Kmax) were 107 ± 16 and 119 ± 7, respectively. The fetal and adult pD2 values for NE-induced increase of [Ca2+]iwere 6.2 ± 0.1 and 6.4 ± 0.1, respectively, whereas maximum [Ca2+]iresponses were 81 ± 9 and 103 ± 15% of Kmax, respectively. After 10−5 M NE-induced contraction, nifedipine resulted in dose-dependent decrease in vessel tone and [Ca2+]iwith pIC50 values for fetal and adult tensions of 7.3 ± 0.1 and 6.6 ± 0.1, respectively ( P < 0.01; n = 4 each), whereas pIC50 for [Ca2+]iresponses were 7.2 ± 0.1 and 6.9 ± 0.1, respectively. The pIC50 values for tension for diltiazem and verapamil were somewhat lower but showed a similar relationship. The receptor-operated Ca2+ channel blocker 2-nitro-4 carboxyphenyl- N,N-diphenyl carbamate showed little effect on NE-induced vessel contractility or [Ca2+]i. In the absence of extracellular Ca2+ for 2 min, 10−5 M NE resulted in markedly attenuated responses of adult MCA tension and [Ca2+]ito 39 ± 7 and 73 ± 8% of control values ( n = 4). For fetal MCA, exposure to extracellular Ca2+concentration resulted in essentially no contractile or [Ca2+]iresponse ( n = 4). Similar blunting of NE-induced tension and [Ca2+]iwas seen in response to 10−3M lanthanum ion. These findings provide evidence to suggest that especially in fetal, but also in adult, ovine MCA, Ca2+ flux via L-type calcium channels plays a key role in NE-induced contraction. In contrast, Ca2+ flux via receptor-operated Ca2+ channels is of less importance. This developmental difference in the role of cerebrovascular plasma membrane Ca2+ channels may be an important association with increased Ca2+sensitivity of the fetal vessels.

Cerebral artery sarcoplasmic reticulum Ca2+ stores and contractility: changes with development

2000 ◽  
Vol 279 (3) ◽  
pp. R860-R873 ◽  
Author(s):  
Wen Long ◽  
Lubo Zhang ◽  
Lawrence D. Longo
Keyword(s):  
Plasma Membrane ◽  
Sarcoplasmic Reticulum ◽  
Cerebral Artery ◽  
Cerebral Arteries ◽  
Induced Responses ◽  
Adult Sheep ◽  
Middle Cerebral Arteries ◽  
Intracellular Ca ◽  
Near Term ◽  
Nonpregnant Adult

To test the hypothesis that sarcoplasmic reticulum (SR) Ca2+ stores play a key role in norepinephrine (NE)-induced contraction of fetal and adult cerebral arteries and that Ca2+ stores change with development, we performed the following study. In main branch middle cerebral arteries (MCA) from near-term fetal (∼140 days) and nonpregnant adult sheep, we measured NE-induced contraction and intracellular Ca2+ concentration ([Ca2+]i) in the absence and presence of different blockers. In adult MCA, after thapsigargin (10−6M), the NE-induced responses of tension and [Ca2+]i were 37 ± 5 and 47 ± 7%, respectively, of control values ( P < 0.01 for each). In the fetal artery, in contrast, this treatment resulted in no significant changes from control. When this was repeated in the absence of extracellular Ca2+, adult MCA increases in tension and [Ca2+]i were 32 ± 5 and 13 ± 3%, respectively, of control. Fetal cerebral arteries, however, showed essentially no response. Ryanodine (RYN, 3 × 10−6 to 10−5 M) resulted in increases in tension and [Ca2+]i in both fetal and adult MCA similar to that seen with NE. For both adult and fetal MCA, the increased tension and [Ca2+]i responses to RYN were essentially eliminated in the presence of zero extracellular Ca2+. These findings provide evidence that in fetal MCA, in contrast to those in the adult, SR Ca2+ stores are of less importance in NE-induced contraction, with such contraction being almost wholly dependent on Ca2+ flux via plasma membrane L-type Ca2+ channels. In addition, they suggest that in both adult and fetal MCA, the RYN receptor is coupled to the plasma membrane Ca2+-activated K+ channel and/or L-type Ca2+ channel.

Cerebral artery KATP- and KCa-channel activity and contractility: changes with development

2000 ◽  
Vol 279 (6) ◽  
pp. R2004-R2014 ◽  
Author(s):  
Wen Long ◽  
Lubo Zhang ◽  
Lawrence D. Longo
Keyword(s):  
Channel Blocker ◽  
Cerebral Arteries ◽  
Dependent Manner ◽  
Channel Activity ◽  
Channel Opener ◽  
Channel Inhibition ◽  
Middle Cerebral Arteries ◽  
Intracellular Ca ◽  
Near Term ◽  
Nonpregnant Adult

The present study was designed to test the hypothesis that in cerebral arteries of the fetus, ATP-sensitive (KATP) and Ca2+-activated K+channels (KCa) play an important role in the regulation of intracellular Ca2+ concentration ([Ca2+]i) and that this differs significantly from that of the adult. In main branch middle cerebral arteries (MCA) from near-term fetal (∼140 days) and nonpregnant adult sheep, simultaneously we measured norepinephrine (NE)-induced responses of vascular tension and [Ca2+]i in the absence and presence of selective K+-channel openers/blockers. In fetal MCA, in a dose-dependent manner, both the KATP-channel opener pinacidil and the KCa-channel opener NS 1619 significantly inhibited NE-induced tension [negative logarithm of the half-maximal inhibitory concentration (pIC50) = 5.0 ± 0.1 and 8.2 ± 0.1, respectively], with a modest decrease of [Ca2+]i. In the adult MCA, in contrast, both pinacidil and NS 1619 produced a significant tension decrease (pIC50 = 5.1 ± 0.1 and 7.6 ± 0.1, respectively) with no change in [Ca2+]i. In addition, the KCa-channel blocker iberiotoxin (10−7 to 10−6 M) resulted in increased tension and [Ca2+]i in both adult and fetal MCA, although the KATP-channel blocker glibenclamide (10−7 to 3 × 10−5 M) failed to do so. Of interest, administration of 10−7 M iberiotoxin totally eliminated vascular contraction and increase in [Ca2+]i seen in response to 10−5M ryanodine. In precontracted fetal cerebral arteries, activation of the KATP and KCa channels significantly decreased both tension and [Ca2+]i, suggesting that both K+ channels play an important role in regulating L-type channel Ca2+ flux and therefore vascular tone in these vessels. In the adult, KATP and the KCa channels also appear to play an important role in this regard; however, in the adult vessel, activation of these channels with resultant vasorelaxation can occur with no significant change in [Ca2+]i. These channels show differing responses to inhibition, e.g., KCa-channel inhibition, resulting in increased tension and [Ca2+]i, whereas KATP-channel inhibition showed no such effect. In addition, the KCa channel appears to be coupled to the sarcoplasmic reticulum ryanodine receptor. Thus differences in plasma membrane K+-channel activity may account, in part, for the differences in the regulation of contractility of fetal and adult cerebral arteries.

Role of iPLA2 and store-operated channels in agonist-induced Ca2+ influx and constriction in cerebral, mesenteric, and carotid arteries

2008 ◽  
Vol 294 (3) ◽  
pp. H1183-H1187 ◽  
Author(s):  
Kristen M. Park ◽  
Mario Trucillo ◽  
Nicolas Serban ◽  
Richard A. Cohen ◽  
Victoria M. Bolotina
Keyword(s):  
Carotid Arteries ◽  
Cerebral Arteries ◽  
Present Evidence ◽  
Voltage Gated ◽  
Functional Inhibition ◽  
Intracellular Ca ◽  
Dependent Pathway ◽  
Secondary Activation

Store-operated channels (SOC) and store-operated Ca2+ entry are known to play a major role in agonist-induced constriction of smooth muscle cells (SMC) in conduit vessels. In microvessels the role of SOC remains uncertain, in as much as voltage-gated L-type Ca2+ (CaL2+) channels are thought to be fully responsible for agonist-induced Ca2+ influx and vasoconstriction. We present evidence that SOC and their activation via a Ca2+-independent phospholipase A2 (iPLA2)-mediated pathway play a crucial role in agonist-induced constriction of cerebral, mesenteric, and carotid arteries. Intracellular Ca2+ in SMC and intraluminal diameter were measured simultaneously in intact pressurized vessels in vitro. We demonstrated that 1) Ca2+ and contractile responses to phenylephrine (PE) in cerebral and carotid arteries were equally abolished by nimodipine (a CaL2+ inhibitor) and 2-aminoethyl diphenylborinate (an inhibitor of SOC), suggesting that SOC and CaL2+ channels may be involved in agonist-induced constriction of cerebral arteries, and 2) functional inhibition of iPLA2β totally inhibited PE-induced Ca2+ influx and constriction in cerebral, mesenteric, and carotid arteries, whereas K+-induced Ca2+ influx and vasoconstriction mediated by CaL2+ channels were not affected. Thus iPLA2-dependent activation of SOC is crucial for agonist-induced Ca2+ influx and vasoconstriction in cerebral, mesenteric, and carotid arteries. We propose that, on PE-induced depletion of Ca2+ stores, nonselective SOC are activated via an iPLA2-dependent pathway and may produce a depolarization of SMC, which could trigger a secondary activation of CaL2+ channels and lead to Ca2+ entry and vasoconstriction.

L-type Ca2+ channels in fetal and adult ovine cerebral arteries

2002 ◽  
Vol 282 (1) ◽  
pp. R131-R138 ◽  
Author(s):  
Arlin B. Blood ◽  
Yu Zhao ◽  
Wen Long ◽  
Lubo Zhang ◽  
Lawrence D. Longo
Keyword(s):  
Carotid Arteries ◽  
Cerebral Arteries ◽  
Bay K 8644 ◽  
Channel Density ◽  
Immunoblotting Assay ◽  
Intracellular Ca ◽  
Common Carotid Arteries ◽  
Fetal Vessels ◽  
Ca2 Channels

Recently, we reported that, whereas in cerebral arteries of the adult a majority of norepinephrine (NE)-induced increase in intracellular Ca2+ concentration ([Ca2+]i) comes from release of the sarcoplasmic reticulum (SR) Ca2+ stores, in the fetus the SR Ca2+ stores are relatively small, and NE-induced increase in [Ca2+]i results mainly from activation of plasma membrane L-type Ca2+ channels (20). In an effort to establish further the role of L-type Ca2+ channels in the developing cerebral arteries, we tested the hypothesis that, in the fetus, increased reliance on plasmalemmal L-type Ca2+ channels is mediated, in part, by increased L-type Ca2+ channel density. We used3H-labeled (+)isopropyl-4-(2,1,3-benzoxadiazol-4-y1)-1,4-dihydro-(2,6-dimethyl-5-methoxycarbonyl)pyridine-3-carboxylate (PN200–110, isradipine) to measure L-type Ca2+ channel density (Bmax) in the cerebral arteries, common carotid artery (CCA), and descending aortae of fetal (∼140 gestation days), newborn (7–10 days), and adult sheep. In the cerebral and common carotid arteries, Bmax values (fmol/mg protein) of fetuses and newborns were significantly greater than those of adults. Western immunoblotting assay also revealed that the density of L-type Ca2+ channel protein in the cerebral arteries and CCA was about twofold greater in the fetus than the adult. Finally, compared with the adult, fetal cerebral arteries demonstrated a significantly greater maximum tension and [Ca2+]i in response to stimulation with the L-type Ca2+ channel agonist Bay K 8644. In addition, Bay K 8644-stimulated fetal vessels demonstrated a maximal tension and [Ca2+]isimilar to that observed in response to stimulation with 10−4 NE. These results support the idea that fetal cerebrovascular smooth muscle relies more on extracellular Ca2+ and L-type Ca2+ channels for contraction than does the adult and that this increased reliance is mediated, in part, by greater L-type Ca2+ channel density. This may have important implications in the regulation of cerebral blood flow in the developing organism.

Release and reloading of intracellular Ca stores after cholinergic stimulation of the parietal cell

1988 ◽  
Vol 254 (4) ◽  
pp. C498-C504 ◽  
Author(s):  
P. A. Negulescu ◽  
T. E. Machen
Keyword(s):  
Plasma Membrane ◽  
Parietal Cell ◽  
Base Line ◽  
Cholinergic Stimulation ◽  
Gastric Glands ◽  
Cholinergic Agonist ◽  
Intracellular Ca ◽  
Lanthanum Ion ◽  
Intact Rabbit ◽  
Stimulation Of

Microspectrofluorimetry was used to measure cytosolic free Ca, Cai, in single parietal cells of intact rabbit gastric glands loaded with the Ca-sensitive fluorescent dye, fura-2. Cells were repeatedly stimulated with the cholinergic agonist carbachol to gain insights into the membrane mechanisms involved in hormonally stimulated Ca metabolism. In either Ca-containing or Ca-free solutions, carbachol (100 microM) caused a rapid (within 30 s) elevation of Cai from a resting level of 100 nM to greater than 600 nM. After the spike, Cai decreased within 3 min to a lower level that was somewhat elevated (greater than 200 nM) over base line. This plateau was dependent on both carbachol and extracellular Ca (Cao) and could be blocked by the addition of atropine (1 microM) or lanthanum ion (La, 50 microM). The spike is due to the release of Ca from internal stores, whereas the plateau is due to Ca entry across the plasma membrane through agonist-controlled, La-inhibitable channels. After a carbachol stimulation of 3 min or longer, reloading of the internal store was absolutely dependent on Cao. Under these conditions, reloading occurred through a La-sensitive (but nifedipine- and verapamil-insensitive) pathway in the plasma membrane. No significant change in Cai was detectable during the reloading. In contrast to the longer treatments, if carbachol stimulation was terminated with atropine while Cai was still elevated, significant reloading occurred from the cytosol.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Regulation of Ca2+sensitization by PKC and rho proteins in ovine cerebral arteries: effects of artery size and age

1998 ◽  
Vol 275 (3) ◽  
pp. H930-H939 ◽  
Author(s):  
Sergey E. Akopov ◽  
Lubo Zhang ◽  
William J. Pearce
Keyword(s):  
G Protein ◽  
Cerebral Arteries ◽  
Rho Kinase ◽  
Cerebrovascular Reactivity ◽  
Rho Proteins ◽  
Experimental Conditions ◽  
Fetal Sheep ◽  
Middle Cerebral Arteries ◽  
Near Term ◽  
Nonpregnant Adult

G protein-regulated Ca2+ sensitivity of vascular contractile proteins plays an important role in cerebrovascular reactivity. The present study examines the intracellular mechanisms that govern G protein-regulated Ca2+ sensitivity in cerebral arteries of different size and age. We studied β-escin-permeabilized segments of common carotid, basilar, and middle cerebral arteries from nonpregnant adult and near-term fetal sheep. Activation of protein kinase C (PKC) by (−)-indolactam V or a phorbol ester produced receptor-independent increases in Ca2+ sensitivity. Such increases were more marked in immature arteries and were inversely correlated with artery size in both mature and immature arteries. However, inhibitors of PKC did not significantly affect increases in Ca2+ sensitivity in responses to either serotonin (5-hydroxytryptamine, 5-HT) or guanosine 5′- O-(3-thiotriphosphate) (GTPγS). Alternatively, deactivation of rho p21, a small G protein associated with Rho kinase, by exotoxin C3 fully prevented increases in Ca2+ sensitivity in responses to 5-HT or GTPγS in both adult and fetal arteries of all types. Neither inhibitors of PKC nor exotoxin C3 altered baseline Ca2+ sensitivity. We conclude that patterns of receptor- and/or G protein-mediated modulation of Ca2+ sensitivity are dependent on an intracellular pathway that involves activation of small G proteins and Rho kinase. In contrast, PKC has little, if any, role in agonist-induced Ca2+ sensitization under the present experimental conditions.

Dual role of PKC in modulating pharmacomechanical coupling in fetal and adult cerebral arteries

2000 ◽  
Vol 279 (4) ◽  
pp. R1419-R1429 ◽  
Author(s):  
Lawrence D. Longo ◽  
Yu Zhao ◽  
Wen Long ◽  
Carolyn Miguel ◽  
Ryan S. Windemuth ◽  
...  
Keyword(s):  
Vascular Tone ◽  
Cerebral Arteries ◽  
Age Groups ◽  
Intracellular Ca ◽  
Membrane Bound ◽  
Near Term ◽  
Fetal Vessel ◽  
Pharmacomechanical Coupling ◽  
Induced Signal ◽  
Two Age Groups

This study tested the hypothesis that protein kinase C (PKC) has dual regulation on norepinephrine (NE)-mediated inositol 1,4,5-trisphosphate [Ins (1,4,5)P3] pathway and vasoconstriction in cerebral arteries from near-term fetal (∼140 gestational days) and adult sheep. Basal PKC activity values (%membrane bound) in fetal and adult cerebral arteries were 38 ± 4% and 32 ± 4%, respectively. In vessels of both age groups, the PKC isoforms α, βI, βII, and δ were relatively abundant. In contrast, compared with the adult, cerebral arteries of the fetus had low levels of PKC-ε. In response to 10−4 M phorbol 12,13-dibutyrate (PDBu; PKC agonist), PKC activity in both fetal and adult cerebral arteries increased 40–50%. After NE stimulation, PKC activation with PDBu exerted negative feedback on Ins(1,4,5)P3 and intracellular Ca2+ concentration ([Ca2+]i) in arteries of both age groups. In turn, PKC inhibition with staurosporine resulted in augmented NE-induced Ins(1,4,5)P3 and [Ca2+]i responses in adult, but not fetal, cerebral arteries. In adult tissues, PKC stimulation by PDBu increased vascular tone, but not [Ca2+]i. In contrast, in the fetal artery, PKC stimulation was associated with an increase in both tone and [Ca2+]i. In the presence of zero extracellular [Ca2+], these PDBu-induced responses were absent in the fetal vessel, whereas they remained unchanged in the adult. We conclude that, although basal PKC activity was similar in fetal and adult cerebral arteries, PKC's role in NE-mediated pharmacomechanical coupling differed significantly in the two age groups. In both fetal and adult cerebral arteries, PKC modulation of NE-induced signal transduction responses would appear to play a significant role in the regulation of vascular tone. The mechanisms differ in the two age groups, however, and this probably relates, in part, to the relative lack of PKC-ε in fetal vessels.

scholarly journals α1-Adrenergic receptor subtype function in fetal and adult cerebral arteries

2010 ◽  
Vol 298 (6) ◽  
pp. H1797-H1806 ◽  
Author(s):  
Ravi Goyal ◽  
Ashwani Mittal ◽  
Nina Chu ◽  
Lubo Zhang ◽  
Lawrence D. Longo
Keyword(s):  
Adrenergic Receptor ◽  
Cerebral Arteries ◽  
Age Groups ◽  
Critical Factor ◽  
Receptor Subtype ◽  
Erk Activation ◽  
Wb 4101 ◽  
Intracellular Ca ◽  
Nonpregnant Adult ◽  
And Function

In the developing fetus, cerebral artery (CA) contractility demonstrates significant functional differences from that of the adult. This may be a consequence of differential activities of α1-adrenergic receptor (α1-AR) subtypes. Thus we tested the hypothesis that maturational differences in adrenergic-mediated CA contractility are, in part, a consequence of differential expression and/or activities of α1-AR subtypes. In CA from fetal (∼140 days) and nonpregnant adult sheep, we used wire myography and imaging, with simultaneous measurement of tension and intracellular Ca2+ concentration ([Ca2+]i), radioimmunoassay, and Western immunoblots to examine phenylephrine (Phe)-induced contractile responses. The α1A-AR antagonists (5-MU and WB-4101) completely inhibited Phe-induced contraction in adult but not fetal CA; however, [Ca2+]i increase was reduced significantly in both age groups. The α1D-AR antagonist (BMY-7378) blocked both Phe-induced contractions and Ca2+ responses to a significantly greater extent in adult compared with fetal CA. In both age groups, inhibition of α1A-AR and α1B-AR, but not α1D-AR, significantly reduced inositol 1,4,5-trisphosphate responses to Phe. Western immunoblots demonstrated that the α1-AR subtype expression was only ∼20% in fetal CA compared with the adult. Moreover, in fetal CA, the α1D-AR was expressed significantly greater than the other two subtypes. Also, in fetal but not adult CA, Phe induced a significant increase in activated ERK1/2; this increase in phosphorylated ERK was blocked by α1B-AR (CEC) and α1D-AR (BMY-7378) inhibitors, but not by α1A-AR inhibitors (5-MU or WB-4101). In conclusion, in the fetal CA, α1B-AR and α1D-AR subtypes play a key role in contractile response as well as in ERK activation. We speculate that in fetal CA α1B-AR and α1D-AR subtypes may be a critical factor associated with cerebrovascular growth and function.

Modeling of Ca2+ flux in pancreatic β-cells: role of the plasma membrane and intracellular stores

2003 ◽  
Vol 285 (1) ◽  
pp. E138-E154 ◽  
Author(s):  
Leonid E. Fridlyand ◽  
Natalia Tamarina ◽  
Louis H. Philipson
Keyword(s):  
Plasma Membrane ◽  
Calcium Oscillations ◽  
Β Cells ◽  
Β Cell ◽  
Pancreatic Β Cells ◽  
Ionic Flux ◽  
Inositol 1,4,5 Trisphosphate ◽  
Intracellular Ca ◽  
Uptake And Release

We have developed a detailed mathematical model of ionic flux in β-cells that includes the most essential channels and pumps in the plasma membrane. This model is coupled to equations describing Ca2+, inositol 1,4,5-trisphosphate (IP3), ATP, and Na+ homeostasis, including the uptake and release of Ca2+ by the endoplasmic reticulum (ER). In our model, metabolically derived ATP activates inward Ca2+ flux by regulation of ATP-sensitive K+ channels and depolarization of the plasma membrane. Results from the simulations support the hypothesis that intracellular Na+ and Ca2+ in the ER can be the main variables driving both fast (2–7 osc/min) and slow intracellular Ca2+ concentration oscillations (0.3–0.9 osc/min) and that the effect of IP3 on Ca2+ leak from the ER contributes to the pattern of slow calcium oscillations. Simulations also show that filling the ER Ca2+ stores leads to faster electrical bursting and Ca2+ oscillations. Specific Ca2+ oscillations in isolated β-cell lines can also be simulated.

Characterizing voltage-dependent Ca2+ channels coupled to VIP release and NO synthesis in enteric synaptosomes

2002 ◽  
Vol 283 (5) ◽  
pp. G1027-G1034 ◽  
Author(s):  
M. Kurjak ◽  
A. Sennefelder ◽  
M. Aigner ◽  
V. Schusdziarra ◽  
H. D. Allescher
Keyword(s):  
Nitric Oxide ◽  
No Synthase ◽  
Channel Blockers ◽  
Voltage Dependent ◽  
Intracellular Ca ◽  
P Type ◽  
Ca2 Channels

In enteric synaptosomes of the rat, the role of voltage-dependent Ca2+channels in K+-induced VIP release and nitric oxide (NO) synthesis was investigated. Basal VIP release was 39 ± 4 pg/mg, and cofactor-substituted NO synthase activity was 7.0 ± 0.8 fmol · mg−1 · min−1. K+ depolarization (65 mM) stimulated VIP release Ca2+ dependently (basal, 100%; K+, 172.2 ± 16.2%; P < 0.05, n = 5). K+-stimulated VIP release was reduced by blockers of the P-type (ω-agatoxin-IVA, 3 × 10−8 M) and N-type (ω-conotoxin-GVIA, 10−6 M) Ca2+ channels by ∼50 and 25%, respectively, but not by blockers of the L-type (isradipine, 10−8 M), Q-type (ω-conotoxin-MVIIC, 10−6 M), or T-type (Ni2+, 10−6 M) Ca2+ channels. In contrast, NO synthesis was suppressed by ω-agatoxin-IVA, ω-conotoxin-GVIA, and isradipine by ∼79, 70, and 70%, respectively, whereas Ni2+ and ω-conotoxin-MVIIC had no effect. These findings are suggestive of a coupling of depolarization-induced VIP release primarily to the P- and N-type Ca2+ channels, whereas NO synthesis is presumably dependent on Ca2+ influx not only via the P- and N- but also via the L-type Ca2+ channel. In contrast, none of the Ca2+ channel blockers affected VIP release evoked by exogenous NO, suggesting that NO induces VIP secretion by a different mechanism, presumably involving intracellular Ca2+ stores.

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