Biology Reference
In-Depth Information
Chapter 44
Assessment of Intracellular Calcium in Cerebral
Artery Myocytes
Matthew A. Nystoriak , Masayo Koide , and George C. Wellman
Abstract
Vascular tone and consequently, cerebral blood fl ow, are regulated by the concentration of global cytosolic
Ca
2+
(global [Ca
2+
]
i
) in vascular smooth muscle. Global [Ca
2+
]
i
is, in part, regulated by discrete Ca
2+
release
events from the sarcoplasmic reticulum (Ca
2+
sparks) which activate plasma membrane large conductance
Ca
2+
-activated K
+
channels to cause membrane potential hyperpolarization, decreased global [Ca
2+
]
i
, and
vasodilation. We have previously observed decreased Ca
2+
spark frequency and elevated global [Ca
2+
]
i
in
cerebral artery myocytes using a rabbit model of subarachnoid hemorrhage. Here, we describe and discuss
laboratory methods and procedures using the fl uorescent Ca
2+
indicator dyes fura-2 and fl uo-4 to assess
global and local Ca
2+
signaling in cerebral arteries and isolated vascular myocytes.
Key words:
Fura-2, Fluo-4, Ca
2+
sparks, Vascular smooth muscle, Subarachnoid hemorrhage
1. Introduction
The cerebral circulation delivers constant blood fl ow to the brain
over a broad range of physiological blood pressures. To achieve
this remarkable phenomenon, cerebral arteries must actively con-
strict to elevations in blood pressure and dilate when blood pres-
sure drops. In cerebral resistance arteries and arterioles, the
constriction (myogenic tone) response to graded increases in intra-
vascular pressure is dependent on membrane potential depolariza-
tion, enhanced Ca
2+
infl ux via L-type voltage-dependent Ca
2+
channels (VDCCs) and elevated global cytosolic Ca
2+
(global
[Ca
2+
]
i
) (
1, 2
). Global [Ca
2+
]
i
represents average Ca
2+
levels
throughout the cytoplasm and mediates contraction of smooth
muscle via Ca
2+
/calmodulin-dependent activation of myosin light
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