Abstract
We developed protocols with intact cultured neonatal rat myocytes to directly evaluate the function of the sarcoplasmic reticulum (SR) Ca-ATPase (or SERCA2), Na-Ca exchange (Na-CaX), and slow Ca transport systems (mitochondria and sarcolemmal Ca-ATPase). Spontaneously heating control cells were compared with cells cultured for 2 days in the presence of verapamil (verapamil-arrested cells VA). Intracellular calcium (Ca(i)) transients were measured by use of indo-1 during (1) spontaneous twitches, (2) contractures induced by rapid application of caffeine (CafC, with and without Na(o)), and (3) twitches induced by brief depolarizations with high [K](o) solution (K- twitches). We also measured mRNA levels for the SR Ca-ATPase and Na-CaX in the same experimental preparations. The t( 1/2 ) for [Ca](i) decline when both the SR Ca uptake and Na-CaX were prevented was the same for control and VA cells (≃20 seconds), indicating unaltered slow Ca transport systems. Similarly, there was no significant difference in the t( 1/2 ) of CafC when Na- CaX was the main mechanism responsible for [Ca](i) decline (t( 1/2 )≃1.5 seconds), indicating unaltered Na-CaX. Conversely, we found nearly a twofold increase in the rate of [Ca](i) decline during K-twitches (control t( 1/2 ), 0.84±0.05 seconds; VA t( 1/2 ), 0.48±0.06 second; P<.001), indicating an increase in SR Ca-pumping activity in VA cells. This was also reflected by a 56% increase in the peak [Ca](i) reached during CafC used to assess maximal SR Ca content (427±49 nmol/L in control versus 665±75 nmol/L in VA cells). In agreement with these functional effects, we found no change in Na-CaX mRNA levels but a marked upregulation of both the SERCA2 mRNA and protein levels in VA cells (to 166±10% and 164±20%, respectively). Thus, verapamil arrest induced an increase in SR Ca uptake (and SERCA2 expression) without affecting the Na-CaX activity (or expression) or the slow Ca transport systems.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 991-997 |
| Number of pages | 7 |
| Journal | Circulation Research |
| Volume | 74 |
| Issue number | 5 |
| State | Published - May 1994 |
| Externally published | Yes |
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Keywords
- Ca(i) transients
- caffeine
- hypertrophy
- Na-Ca exchange
- rat
- SR Ca-ATPase
ASJC Scopus subject areas
- Physiology
- Cardiology and Cardiovascular Medicine
Cite this
Contractile arrest increases sarcoplasmic reticulum calcium uptake and SERCA2 gene expression in cultured neonatal rat heart cells. / Bassani, José W M; Qi, Ming; Samarel, Allen M.; Bers, Donald M.
In: Circulation Research, Vol. 74, No. 5, 05.1994, p. 991-997.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Contractile arrest increases sarcoplasmic reticulum calcium uptake and SERCA2 gene expression in cultured neonatal rat heart cells
AU - Bassani, José W M
AU - Qi, Ming
AU - Samarel, Allen M.
AU - Bers, Donald M
PY - 1994/5
Y1 - 1994/5
N2 - We developed protocols with intact cultured neonatal rat myocytes to directly evaluate the function of the sarcoplasmic reticulum (SR) Ca-ATPase (or SERCA2), Na-Ca exchange (Na-CaX), and slow Ca transport systems (mitochondria and sarcolemmal Ca-ATPase). Spontaneously heating control cells were compared with cells cultured for 2 days in the presence of verapamil (verapamil-arrested cells VA). Intracellular calcium (Ca(i)) transients were measured by use of indo-1 during (1) spontaneous twitches, (2) contractures induced by rapid application of caffeine (CafC, with and without Na(o)), and (3) twitches induced by brief depolarizations with high [K](o) solution (K- twitches). We also measured mRNA levels for the SR Ca-ATPase and Na-CaX in the same experimental preparations. The t( 1/2 ) for [Ca](i) decline when both the SR Ca uptake and Na-CaX were prevented was the same for control and VA cells (≃20 seconds), indicating unaltered slow Ca transport systems. Similarly, there was no significant difference in the t( 1/2 ) of CafC when Na- CaX was the main mechanism responsible for [Ca](i) decline (t( 1/2 )≃1.5 seconds), indicating unaltered Na-CaX. Conversely, we found nearly a twofold increase in the rate of [Ca](i) decline during K-twitches (control t( 1/2 ), 0.84±0.05 seconds; VA t( 1/2 ), 0.48±0.06 second; P<.001), indicating an increase in SR Ca-pumping activity in VA cells. This was also reflected by a 56% increase in the peak [Ca](i) reached during CafC used to assess maximal SR Ca content (427±49 nmol/L in control versus 665±75 nmol/L in VA cells). In agreement with these functional effects, we found no change in Na-CaX mRNA levels but a marked upregulation of both the SERCA2 mRNA and protein levels in VA cells (to 166±10% and 164±20%, respectively). Thus, verapamil arrest induced an increase in SR Ca uptake (and SERCA2 expression) without affecting the Na-CaX activity (or expression) or the slow Ca transport systems.
AB - We developed protocols with intact cultured neonatal rat myocytes to directly evaluate the function of the sarcoplasmic reticulum (SR) Ca-ATPase (or SERCA2), Na-Ca exchange (Na-CaX), and slow Ca transport systems (mitochondria and sarcolemmal Ca-ATPase). Spontaneously heating control cells were compared with cells cultured for 2 days in the presence of verapamil (verapamil-arrested cells VA). Intracellular calcium (Ca(i)) transients were measured by use of indo-1 during (1) spontaneous twitches, (2) contractures induced by rapid application of caffeine (CafC, with and without Na(o)), and (3) twitches induced by brief depolarizations with high [K](o) solution (K- twitches). We also measured mRNA levels for the SR Ca-ATPase and Na-CaX in the same experimental preparations. The t( 1/2 ) for [Ca](i) decline when both the SR Ca uptake and Na-CaX were prevented was the same for control and VA cells (≃20 seconds), indicating unaltered slow Ca transport systems. Similarly, there was no significant difference in the t( 1/2 ) of CafC when Na- CaX was the main mechanism responsible for [Ca](i) decline (t( 1/2 )≃1.5 seconds), indicating unaltered Na-CaX. Conversely, we found nearly a twofold increase in the rate of [Ca](i) decline during K-twitches (control t( 1/2 ), 0.84±0.05 seconds; VA t( 1/2 ), 0.48±0.06 second; P<.001), indicating an increase in SR Ca-pumping activity in VA cells. This was also reflected by a 56% increase in the peak [Ca](i) reached during CafC used to assess maximal SR Ca content (427±49 nmol/L in control versus 665±75 nmol/L in VA cells). In agreement with these functional effects, we found no change in Na-CaX mRNA levels but a marked upregulation of both the SERCA2 mRNA and protein levels in VA cells (to 166±10% and 164±20%, respectively). Thus, verapamil arrest induced an increase in SR Ca uptake (and SERCA2 expression) without affecting the Na-CaX activity (or expression) or the slow Ca transport systems.
KW - Ca(i) transients
KW - caffeine
KW - hypertrophy
KW - Na-Ca exchange
KW - rat
KW - SR Ca-ATPase
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UR - http://www.scopus.com/inward/citedby.url?scp=0028274023&partnerID=8YFLogxK
M3 - Article
C2 - 8156646
AN - SCOPUS:0028274023
VL - 74
SP - 991
EP - 997
JO - Circulation Research
JF - Circulation Research
SN - 0009-7330
IS - 5
ER -