Bradykinin increases permeability by calcium and 5-lipoxygenase in the ECV304/C6 cell culture model of the blood-brain barrier

A S Easton; N J Abbott

doi:10.1016/S0006-8993(02)03281-X

Bradykinin increases permeability by calcium and 5-lipoxygenase in the ECV304/C6 cell culture model of the blood-brain barrier

A S Easton, N J Abbott

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Abstract

The blood-brain barrier (BBB) was modelled in this study using ECV304 cells in co-culture with rat C6 glioma cells, which resulted in elevated transendothelial electrical resistance (TEER). The inflammatory mediator bradykinin (1 muM) was studied and found to induce a fall in TEER; the link between this change and intracellular free calcium concentration ([Ca2+](i)) was then examined. 1 muM bradykinin produced a peak-plateau increase in [Ca2+](i). The peak showed desensitization and was dose dependent (over 0.1 nM to 1 muM). The [Ca2+](i) increase was blocked by the B-2 antagonist HOE 140 (1 muM) without effect from a B-1 agonist and antagonist. The plateau response was abolished in Ca2+-free solution containing 2 mM EDTA, and also by the Ca2+ channel blockers lanthanum, La3+ (10 muM), and SKF 96365 (100 muM). The store Ca(2+)ATPase inhibitor thapsigargin (1 muM) abolished the peak response. The putative phospholipase C inhibitors, U73122 (20 muM) and ETH-18-OCH3 (100 muM), unexpectedly increased [Ca2+](i); after their application, bradykinin was ineffective. Agents without effect on Ca2+ responses to bradykinin included the phospholipase A(2) (PLA(2)) inhibitor aristolochic acid (0.5 mM), cyclooxygenase inhibitor indomethacin (100 muM), 5-lipoxygenase inhibitor nordihydroguaiaretic acid, NDGA (100 P M), calphostin C (0.5 muM), L-NAME (I mM) and nifedipine (10 muM). The fall in TEER from bradykinin was blocked by HOE 140 U73122 and thapsigargin combined with La3+, and also by aristolochic acid and NDGA, but not indomethacin, calphostin C Or L-NAME. U73122 increased TEER while ETH-18-OCH3 reduced it. Thus bradykinin reduced TEER through B-2 receptor-linked release of Ca2+ from thapsigargin-sensitive stores, leading to activation of PLA(2) and metabolism of arachidonic acid by 5-lipoxygenase. (C) 2002 Elsevier Science B.V. All rights reserved.

Original language	English
Pages (from-to)	157 - 169
Number of pages	13
Journal	Brain Research
Volume	953
Issue number	1-2
DOIs	https://doi.org/10.1016/S0006-8993(02)03281-X
Publication status	Published - 20 Nov 2002

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10.1016/S0006-8993(02)03281-X

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@article{eac67deb405b42799a5c8bc2b132a618,

title = "Bradykinin increases permeability by calcium and 5-lipoxygenase in the ECV304/C6 cell culture model of the blood-brain barrier",

abstract = "The blood-brain barrier (BBB) was modelled in this study using ECV304 cells in co-culture with rat C6 glioma cells, which resulted in elevated transendothelial electrical resistance (TEER). The inflammatory mediator bradykinin (1 muM) was studied and found to induce a fall in TEER; the link between this change and intracellular free calcium concentration ([Ca2+](i)) was then examined. 1 muM bradykinin produced a peak-plateau increase in [Ca2+](i). The peak showed desensitization and was dose dependent (over 0.1 nM to 1 muM). The [Ca2+](i) increase was blocked by the B-2 antagonist HOE 140 (1 muM) without effect from a B-1 agonist and antagonist. The plateau response was abolished in Ca2+-free solution containing 2 mM EDTA, and also by the Ca2+ channel blockers lanthanum, La3+ (10 muM), and SKF 96365 (100 muM). The store Ca(2+)ATPase inhibitor thapsigargin (1 muM) abolished the peak response. The putative phospholipase C inhibitors, U73122 (20 muM) and ETH-18-OCH3 (100 muM), unexpectedly increased [Ca2+](i); after their application, bradykinin was ineffective. Agents without effect on Ca2+ responses to bradykinin included the phospholipase A(2) (PLA(2)) inhibitor aristolochic acid (0.5 mM), cyclooxygenase inhibitor indomethacin (100 muM), 5-lipoxygenase inhibitor nordihydroguaiaretic acid, NDGA (100 P M), calphostin C (0.5 muM), L-NAME (I mM) and nifedipine (10 muM). The fall in TEER from bradykinin was blocked by HOE 140 U73122 and thapsigargin combined with La3+, and also by aristolochic acid and NDGA, but not indomethacin, calphostin C Or L-NAME. U73122 increased TEER while ETH-18-OCH3 reduced it. Thus bradykinin reduced TEER through B-2 receptor-linked release of Ca2+ from thapsigargin-sensitive stores, leading to activation of PLA(2) and metabolism of arachidonic acid by 5-lipoxygenase. (C) 2002 Elsevier Science B.V. All rights reserved.",

author = "Easton, {A S} and Abbott, {N J}",

year = "2002",

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doi = "10.1016/S0006-8993(02)03281-X",

language = "English",

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T1 - Bradykinin increases permeability by calcium and 5-lipoxygenase in the ECV304/C6 cell culture model of the blood-brain barrier

AU - Easton, A S

AU - Abbott, N J

PY - 2002/11/20

Y1 - 2002/11/20

N2 - The blood-brain barrier (BBB) was modelled in this study using ECV304 cells in co-culture with rat C6 glioma cells, which resulted in elevated transendothelial electrical resistance (TEER). The inflammatory mediator bradykinin (1 muM) was studied and found to induce a fall in TEER; the link between this change and intracellular free calcium concentration ([Ca2+](i)) was then examined. 1 muM bradykinin produced a peak-plateau increase in [Ca2+](i). The peak showed desensitization and was dose dependent (over 0.1 nM to 1 muM). The [Ca2+](i) increase was blocked by the B-2 antagonist HOE 140 (1 muM) without effect from a B-1 agonist and antagonist. The plateau response was abolished in Ca2+-free solution containing 2 mM EDTA, and also by the Ca2+ channel blockers lanthanum, La3+ (10 muM), and SKF 96365 (100 muM). The store Ca(2+)ATPase inhibitor thapsigargin (1 muM) abolished the peak response. The putative phospholipase C inhibitors, U73122 (20 muM) and ETH-18-OCH3 (100 muM), unexpectedly increased [Ca2+](i); after their application, bradykinin was ineffective. Agents without effect on Ca2+ responses to bradykinin included the phospholipase A(2) (PLA(2)) inhibitor aristolochic acid (0.5 mM), cyclooxygenase inhibitor indomethacin (100 muM), 5-lipoxygenase inhibitor nordihydroguaiaretic acid, NDGA (100 P M), calphostin C (0.5 muM), L-NAME (I mM) and nifedipine (10 muM). The fall in TEER from bradykinin was blocked by HOE 140 U73122 and thapsigargin combined with La3+, and also by aristolochic acid and NDGA, but not indomethacin, calphostin C Or L-NAME. U73122 increased TEER while ETH-18-OCH3 reduced it. Thus bradykinin reduced TEER through B-2 receptor-linked release of Ca2+ from thapsigargin-sensitive stores, leading to activation of PLA(2) and metabolism of arachidonic acid by 5-lipoxygenase. (C) 2002 Elsevier Science B.V. All rights reserved.

AB - The blood-brain barrier (BBB) was modelled in this study using ECV304 cells in co-culture with rat C6 glioma cells, which resulted in elevated transendothelial electrical resistance (TEER). The inflammatory mediator bradykinin (1 muM) was studied and found to induce a fall in TEER; the link between this change and intracellular free calcium concentration ([Ca2+](i)) was then examined. 1 muM bradykinin produced a peak-plateau increase in [Ca2+](i). The peak showed desensitization and was dose dependent (over 0.1 nM to 1 muM). The [Ca2+](i) increase was blocked by the B-2 antagonist HOE 140 (1 muM) without effect from a B-1 agonist and antagonist. The plateau response was abolished in Ca2+-free solution containing 2 mM EDTA, and also by the Ca2+ channel blockers lanthanum, La3+ (10 muM), and SKF 96365 (100 muM). The store Ca(2+)ATPase inhibitor thapsigargin (1 muM) abolished the peak response. The putative phospholipase C inhibitors, U73122 (20 muM) and ETH-18-OCH3 (100 muM), unexpectedly increased [Ca2+](i); after their application, bradykinin was ineffective. Agents without effect on Ca2+ responses to bradykinin included the phospholipase A(2) (PLA(2)) inhibitor aristolochic acid (0.5 mM), cyclooxygenase inhibitor indomethacin (100 muM), 5-lipoxygenase inhibitor nordihydroguaiaretic acid, NDGA (100 P M), calphostin C (0.5 muM), L-NAME (I mM) and nifedipine (10 muM). The fall in TEER from bradykinin was blocked by HOE 140 U73122 and thapsigargin combined with La3+, and also by aristolochic acid and NDGA, but not indomethacin, calphostin C Or L-NAME. U73122 increased TEER while ETH-18-OCH3 reduced it. Thus bradykinin reduced TEER through B-2 receptor-linked release of Ca2+ from thapsigargin-sensitive stores, leading to activation of PLA(2) and metabolism of arachidonic acid by 5-lipoxygenase. (C) 2002 Elsevier Science B.V. All rights reserved.

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U2 - 10.1016/S0006-8993(02)03281-X

DO - 10.1016/S0006-8993(02)03281-X

M3 - Article

SN - 1873-2747

VL - 953

SP - 157

EP - 169

JO - Brain Research

JF - Brain Research

IS - 1-2

ER -

Bradykinin increases permeability by calcium and 5-lipoxygenase in the ECV304/C6 cell culture model of the blood-brain barrier

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