Calcium influx pathways in rat CNS pericytes.
In central nervous system (CNS), pericytes have been proposed to play a role in broad functional activities including blood-brain barrier, microcirculation, and macrophage activity. However, contractile responses and Ca2+ signaling in CNS pericytes have not been elucidated. The aim of the present study is to investigate contractility and Ca2+ influx pathway in CNS pericytes. CNS pericytes were cultured from rat brain. Contraction of the pericytes in response to various stimuli was evaluated by the change in surface area measured by a light microscope with a digital camera. Reverse transcription and polymerase chain reaction (RT-PCR) was performed to examine the expression of mRNA of alpha-smooth muscle actin. Intracellular Ca2+ was measured using fura-2 fluorescence spectroscopy. A23187 (Ca2+ ionophore), high external K+ (4 x 10(-2) mol/l), endothelin-1, and serotonin induced contraction of CNS pericytes. RT-PCR analysis revealed the expression of alpha-smooth muscle actin in CNS pericytes. Cytosolic Ca2+ ([Ca2+]i) increased after application of high concentration of external K+, tetraethylammonium, and charybdotoxin, which was inhibited by nicardipine and removal of external Ca2+. Angiotensin-II, serotonin, acetylcholine, ATP, and endothelin-1 caused biphasic response in [Ca2+]i. In response to these agents, [Ca2+]i rapidly increased and then decayed to a relatively constant Ca2+ plateau. The Ca2+ plateau was partially inhibited by nicardipine and completely abolished by omission of external Ca2+. After intracellular Ca2+ store was depleted by the removal of external Ca2+ and addition of thapsigargin, reapplication of external Ca2+ evoked increases in [Ca2+]i. These results indicate that CNS pericytes express mRNA of alpha-smooth muscle actin and possess contractile ability. In CNS pericytes, resting membrane potential is regulated by large conductance Ca2+-activated K+ channels and Ca2+ enters into the cells via L-type voltage-dependent Ca2+ channels, agonist-activated Ca2+ permeable channels, and capacitative Ca2+ entry pathways.
Kamouchi
Masahiro
M
Department of Medicine and Clinical Science, Graduate School of Medical Sciences, Kyushu University, Maidashi 3-1-1, Higashi, Fukuoka 812-8582, Japan. kamouchi@intmed2.med.kyushu-u.ac.jp
Kitazono
Takanari
T
Ago
Tetsuro
T
Wakisaka
Masanori
M
Ooboshi
Hiroaki
H
Ibayashi
Setsuro
S
Iida
Mitsuo
M
D000109:Acetylcholine / Q000378:metabolism; D000199:Actins / Q000235:genetics / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D020013:Calcium Signaling / Q000502:physiology*; D002465:Cell Movement / Q000187:drug effects; D002478:Cells, Cultured; D002490:Central Nervous System / Q000166:cytology* / Q000378:metabolism; D004343:Drug Implants; D019332:Endothelin-1 / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005110:Extracellular Space / Q000187:drug effects / Q000378:metabolism; D016257:Fura-2 / Q000378:metabolism; D007476:Ionophores / Q000494:pharmacology; D008297:Male; D009529:Nicardipine / Q000494:pharmacology; D020286:Pericytes / Q000187:drug effects / Q000378:metabolism*; D011188:Potassium / Q000494:pharmacology; D026902:Potassium Channel Blockers / Q000494:pharmacology; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D012701:Serotonin / Q000494:pharmacology; D019789:Tetraethylammonium / Q000494:pharmacology
D000199:Actins; D002121:Calcium Channel Blockers; D004343:Drug Implants; D019332:Endothelin-1; D004791:Enzyme Inhibitors; D007476:Ionophores; D026902:Potassium Channel Blockers; D012333:RNA, Messenger; D012701:Serotonin; D000001:Calcimycin; D019789:Tetraethylammonium; D000255:Adenosine Triphosphate; D009529:Nicardipine; D000109:Acetylcholine; D011188:Potassium; D002118:Calcium; D016257:Fura-2