Ryanodine receptors regulate arterial diameter and wall [Ca2+] in cerebral arteries of rat via Ca2+-dependent K+ channels - PubMed (original) (raw)

Ryanodine receptors regulate arterial diameter and wall [Ca2+] in cerebral arteries of rat via Ca2+-dependent K+ channels

H J Knot et al. J Physiol. 1998.

Abstract

1. The effects of inhibitors of ryanodine-sensitive calcium release (RyR) channels in the sarcoplasmic reticulum (SR) and Ca2+-dependent potassium (KCa) channels on the membrane potential, intracellular [Ca2+], and diameters of small pressurized (60 mmHg) cerebral arteries (100-200 micron) were studied using digital fluorescence video imaging of arterial diameter and wall [Ca2+], combined with microelectrode measurements of arterial membrane potential. 2. Ryanodine (10 microM), an inhibitor of RyR channels, depolarized by 9 mV, increased intracellular [Ca2+] by 46 nM and constricted pressurized (to 60 mmHg) arteries with myogenic tone by 44 micron (approximately 22 %). Iberiotoxin (100 nM), a blocker of KCa channels, under the same conditions, depolarized the arteries by 10 mV, increased arterial wall calcium by 51 nM, and constricted by 37 micron (approximately 19 %). The effects of ryanodine and iberiotoxin were not additive and were blocked by inhibitors of voltage-dependent Ca2+ channels. 3. Caffeine (10 mM), an activator of RyR channels, transiently increased arterial wall [Ca2+] by 136 +/- 9 nM in control arteries and by 158 +/- 12 nM in the presence of iberiotoxin. Caffeine was relatively ineffective in the presence of ryanodine, increasing [calcium] by 18 +/- 5 nM. 4. In the presence of blockers of voltage-dependent Ca2+ channels (nimodipine, diltiazem), ryanodine and inhibitors of the SR calcium ATPase (thapsigargin, cyclopiazonic acid) were without effect on arterial wall [Ca2+] and diameter. 5. These results suggest that local Ca2+ release originating from RyR channels (Ca2+ sparks) in the SR of arterial smooth muscle regulates myogenic tone in cerebral arteries solely through activation of KCa channels, which regulate membrane potential through tonic hyperpolarization, thus limiting Ca2+ entry through L-type voltage-dependent Ca2+ channels. KCa channels therefore act as a negative feedback control element regulating arterial diameter through a reduction in global intracellular free [Ca2+].

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Figures

Figure 8

Figure 8. Proposed scheme for the regulation of steady-state arterial wall calcium and diameter by ryanodine-sensitive Ca2+-release channels in the sarcoplasmic reticulum

Graded increase of intravascular pressure (10–100 mmHg) depolarizes the smooth muscle cells in the arterial wall (from -63 to −36 mV; Knot & Nelson, 1998), which increases the steady-state open-state probability of L-type voltage-dependent Ca2+ channels (12-fold increase in steady-state open probability from -50 to −30 mV; Rubart et al. 1996). This leads to an increase in steady Ca2+ influx through voltage-dependent Ca2+ channels to elevate steady-state arterial wall [Ca2+] (from 119 to 245 n

m

; Knot & Nelson, 1998). Increased arterial wall [Ca2+] activates myosin light chain kinase, which leads to steady force development, and maintained constriction (‘myogenic tone’). The depolarization-induced increase in steady-state calcium channel activity would increase RyR open probability (‘calcium spark’ frequency) through changes in local and global cytoplasmic calcium (Cannell et al. 1995; Santana et al. 1996), as well as through an elevation of SR calcium load. Increased calcium spark frequency would increase the activity of KCa channels (increased STOC (spontaneous transient outward current) frequency) (Nelson et al. 1995). This proposed mechanism accounts for our previous observations on the regulation of arterial diameter by KCa channels (Brayden & Nelson, 1992; Knot & Nelson, 1995) and their regulation by Ca2+ sparks (Fig. 4, Nelson et al. 1995), and is supported by our new results in this study using direct measurements of arterial membrane potential, arterial wall [Ca2+] and diameter. PLB, phospholamban.

Figure 2

Figure 2. Iberiotoxin increases pressure-induced constrictions

Elevation of intravascular pressure above 10 mmHg causes a sustained decrease in arterial diameter (‘myogenic tone’) (•) from the diameters measured in the absence of external calcium (^). The dihydropyridine calcium channel inhibitor nimodipine (Nimod; 10 n

m

; ▵) dilated the pressurized arteries to steady-state diameters observed in the absence of external calcium. Iberiotoxin caused a sustained constriction above 40 mmHg (▪), but it did not alter the diameters of arteries that had been dilated by nimodipine (□) (_n_= 7). * Significant difference compared with control conditions, P < 0.05.

Figure 3

Figure 3. Ryanodine increases arterial wall Ca2+ and constricts pressurized cerebral arteries

Original recordings of the simultaneous measurement of arterial wall [Ca2+] and diameter in pressurized (to 60 mmHg) posterior cerebral arteries loaded with fura-2. Elevation of intravascular pressure from 10 to 60 mmHg caused a sustained elevation in arterial wall calcium and a sustained constriction relative to Ca2+-free PSS. Ryanodine and high potassium (61 m

m

) increased arterial wall [Ca2+] (A) and decreased arterial diameter (B). Iberiotoxin was without effect in the presence of ryanodine. The dihydropyridine calcium channel inhibitor nimodipine decreased arterial wall calcium in the presence of ryanodine and iberiotoxin. The level of arterial wall calcium at 60 mmHg in the absence of drugs or high potassium is indicated by the horizontal dotted line. The changes (means ±

s.e.m.

, _n_= 6) in arterial wall calcium (C) and diameter (D) in response to high potassium (61 m

m

), ryanodine, ryanodine + iberiotoxin, ryanodine + iberiotoxin + high potassium, and ryanodine + iberiotoxin + high potassium + nimodipine are summarized. Ryanodine and iberiotoxin did not have an effect in the presence of high potassium, consistent with these agents acting on potassium channels. Iberiotoxin was ineffective in the presence of ryanodine, suggesting that calcium released by ryanodine receptors is the primary calcium that activates KCa channels. Nimodipine prevented the effects of high potassium, iberiotoxin and ryanodine, consistent with iberiotoxin and ryanodine requiring functional voltage-dependent calcium channels to have an effect.

Figure 4

Figure 4. Iberiotoxin increases arterial wall Ca2+ and constricts pressurized cerebral arteries

Original recordings of the simultaneous measurement of arterial wall [Ca2+] and diameter in pressurized (to 60 mmHg) posterior cerebral arteries loaded with fura-2. Elevation of intravascular pressure from 10 to 60 mmHg caused a sustained elevation in arterial wall [calcium] and a sustained constriction. Iberiotoxin (100 n

m

) increased arterial wall [Ca2+] (A) and decreased arterial diameter (B). Ryanodine was without effect in the presence of iberiotoxin. High potassium (61 m

m

) elevated arterial wall calcium and decreased arterial diameter in the presence of iberiotoxin and ryanodine. The dihydropyridine calcium channel inhibitor nimodipine decreased arterial wall calcium in the presence of ryanodine, iberiotoxin and high potassium. The level of arterial wall calcium at 60 mmHg in the absence of drugs or high potassium is indicated by the horizontal dotted line. The changes (means ±

s.e.m.

, _n_= 6, 7) in arterial wall calcium (C) and diameter (D) in response to high potassium (61 m

m

), iberiotoxin, ryanodine + iberiotoxin, ryanodine + iberiotoxin + high potassium, and ryanodine + iberiotoxin + high potassium + nimodipine are summarized. Ryanodine and iberiotoxin did not have an effect in the presence of high potassium, consistent with these agents acting on potassium channels. In the presence of iberiotoxin, ryanodine was without effect on steady-state arterial wall calcium and diameter, suggesting that in the steady state, ryanodine receptors act to regulate arterial wall calcium and diameter through KCa channels. Nimodipine prevented the effects of high potassium, iberiotoxin and ryanodine, consistent with iberiotoxin and ryanodine requiring functional voltage-dependent calcium channels to have an effect.

Figure 1

Figure 1. Inhibitors of ryanodine-sensitive calcium release channels (ryanodine) and KCa channels (iberiotoxin) depolarize pressurized cerebral arteries

A, ryanodine (10 μ

m

) depolarizes the membrane potential of a pressurized (to 60 mmHg) cerebral artery, and iberiotoxin (IbTx; 100 n

m

) has no effect in the presence of ryanodine. B, iberiotoxin depolarizes the membrane potential of a pressurized (to 60 mmHg) cerebral artery, and ryanodine has no effect in the presence of iberiotoxin. C, ryanodine (RYA)- and iberiotoxin-induced depolarizations of pressurized cerebral arteries are similar and not additive. The depolarization of these arteries to raising extracellular K+ from 6 to 61 m

m

(61K) is illustrated. Data are represented as means ±

s.d.

(_n_= 3–10 arteries as indicated by the number above the bars).

Figure 5

Figure 5. The effects of ryanodine and iberiotoxin on arterial Ca2+ and diameter are mediated via changes in membrane potential

Arterial diameter and wall [Ca2+] in the presence of 10 μ

m

ryanodine and 100 n

m

iberiotoxin, alone or together are compared with the predicted diameter and [Ca2+] of a pressurized (60 mmHg) cerebral artery that was depolarized by 9 mV (from Fig. 9 in Knot & Nelson, 1998). A, bar graph summarizing the data from 6–7 pressurized (60 mmHg) arteries. There is no significant difference between any of these conditions (for P < 0.05). B, the effects of ryanodine (▵) and iberiotoxin (^) alone and in the presence of 61 m

m

K+ and 10 n

m

nimodipine are plotted on the Ca2+-diameter relationship predicted by changing arterial membrane potential (data points from protocols used in Figs 3 and 4). The continuous line is the predicted relationship based on measured data from Fig. 10 in Knot & Nelson (1998).

Figure 6

Figure 6. Effect of ryanodine and iberiotoxin on arterial wall [Ca2+] and caffeine-induced Ca2+ release from the sarcoplasmic reticulum in pressurized cerebral arteries with and without endothelium

A, the peak changes in arterial wall calcium in response to a bolus application of 10 m

m

caffeine (< 3 s total bath turnover) in pressurized (to 60 mmHg) arteries (with and without endothelium), bathed in iberiotoxin (100 n

m

) or ryanodine (10 μ

m

), are shown. B, removal of the endothelium did not alter steady-state arterial wall [Ca2+] of pressurized (to 60 mmHg) arteries, in the absence and presence of iberiotoxin and ryanodine. *P < 0.05, †P < 0.02, compared with control (PSS).

Figure 7

Figure 7. Ryanodine and thapsigargin do not increase arterial wall [calcium] or decrease arterial diameter in the presence of L-type Ca2+ channel inhibitors

A, original trace showing the effect of 10 μ

m

ryanodine and 100 n

m

thapsigargin (Thg) on arterial wall Ca2+ in pressurized (to 60 mmHg) cerebral arteries in the presence of 30 μ

m

diltiazem. B, original trace (different artery from that shown in A) illustrating the effect of 10 μ

m

ryanodine (in superfusate), 100 n

m

iberiotoxin (bolus into the bath), and thapsigargin (in superfusate) on arterial diameter in the presence of 100 n

m

nimodipine.

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