Relationship between SR Ca2+ release and OI

We have reported previously that TT organization declines in individual cardiac myocytes in intact hypertensive rat hearts and that the number of myocytes showing poor organization increases as myocardial mechanics show signs first of diastolic and then of systolic HF (Aistrup et al. 2013; Shah et al. 2013). Because of the critical nature of the fundamental relationship between cell architecture in the form of TTs and activation of SR Ca2+ release during normal EC coupling, there has been a great deal of interest in how TT remodeling affects Ca2+ transients, particularly SR Ca2+ release. Figure 2 (Panel I) shows a 2‐D image and the corresponding linescans for a myocyte with high TT organization. Figure 2IA shows a 2‐D image of a myocyte from a 7‐month‐old WKY with a high OI value (0.84). The white line in the 2‐D image represents the location in which Ca2+ transient measurements were recorded along the length of the cell in subsequent panels. Figure 2IB shows the corresponding linescan image obtained during basal pacing at a relatively low BCL of 700 msec, with the average intensity profile for the transients located immediately above the image. This linescan shows a set of transients in which the individual transients at all sarcomeres are consistent and all rise rapidly followed by a gradual decay during removal of Ca2+ from the cytoplasm. The white vertical line indicates the initiation time for each transient and the fact that there is virtually no deviation in fluorescence intensity along the line indicates the uniformity of release along the cell length. Since we would expect that any heterogeneities in SR Ca2+ release would be exaggerated during rapid pacing, we also repeated the pacing protocol at a fast rate in this heart (BCL = 300 msec, Figure 2IC). Virtually identical results were obtained during rapid pacing where synchronous and rapid activation of the transient was achieved along the entire cell length. The three cellular regions (indicated as R1, R2, and R3 by the dashed red lines) show Ca2+ transients of both small and large magnitude in which rise time is rapid and uniform throughout the cell that is independent of magnitude and location.

Figure 2 Open in figure viewer PowerPoint 2+ transients (B–C) at BCL = 700 and 300 msec, respectively. Top tracing shows the mean intensity along each line shown in the linescan image below. Three regions (dotted red lines: R1, R2, and R3) are indicated on this image showing representative transients with rapid Ca2+ release. The red horizontal dashed lines have been positioned at 50% of peak for both basal and rapid pacing rates where TR50 is measured. Vertical white lines are placed at the initiation of each transient. Panel II. TTs and Ca2+ transients in a myocyte with severe TT loss from a 12‐month‐old SHR. (A) 2‐D image of a myocyte with poor organization. The white line indicates the position of the scan line. (B–C) corresponding Ca2+ transients. Three representative regions (dotted red lines: R1, R2, and R3) are indicated from this image showing representative transients with varying rates of slow Ca2+ release. The lines have been positioned at the 50% of peak for both basal and rapid pacing rates. The red horizontal dashed lines have been positioned at 50% of peak for both basal and rapid pacing rates where TR 50 is measured. Vertical white lines are placed at the initiation of each transient. Panel III: Summary of the relationships between TR 50 HI and OI during basal (A) and rapid (B) pacing for SHRs and WKY rats (depicted by filled and empty circles, respectively). The inset shows the Ca2+ release in two different regions (R1 and R2) of a myocyte from a 12 month SHR similar to that in Figure N = 4 WKY hearts and 12 SHR hearts. Panel I: Representative 2‐D and linescan images for cells with high value of OI from a 7‐month‐old WKY. (A) 2‐D image of a well‐organized myocyte. The white line indicates the position of the scan line. (B–C) corresponding linescan images showing Catransients (B–C) at BCL = 700 and 300 msec, respectively. Top tracing shows the mean intensity along each line shown in the linescan image below. Three regions (dotted red lines: R1, R2, and R3) are indicated on this image showing representative transients with rapid Carelease. The red horizontal dashed lines have been positioned at 50% of peak for both basal and rapid pacing rates where TR50 is measured. Vertical white lines are placed at the initiation of each transient. Panel IITTs and Catransients in a myocyte with severe TT loss from a 12‐month‐old SHR. (A) 2‐D image of a myocyte with poor organization. The white line indicates the position of the scan line. (B–C) corresponding Catransients. Three representative regions (dotted red lines: R1, R2, and R3) are indicated from this image showing representative transients with varying rates of slow Carelease. The lines have been positioned at the 50% of peak for both basal and rapid pacing rates. The red horizontal dashed lines have been positioned at 50% of peak for both basal and rapid pacing rates where TRis measured. Vertical white lines are placed at the initiation of each transient. Panel III: Summary of the relationships between TRHI and OI during basal (A) and rapid (B) pacing for SHRs and WKY rats (depicted by filled and empty circles, respectively). The inset shows the Carelease in two different regions (R1 and R2) of a myocyte from a 12 month SHR similar to that in Figure 3 with the time of TR50) indicated by the vertical arrows.= 4 WKY hearts and 12 SHR hearts.

Figure 2 (Panel IIA) shows a myocyte from a 12‐month‐old SHR with poor TT organization and a highly disrupted TT network comprised of a few scattered TTs (OI = 0.64). Figure 2IIB shows the corresponding linescan image during basal pacing with a small and variable Ca2+ cycling along the length of the cell as indicated by the variability in fluorescence intensity to the right of the white line indicating the time of initiation. Heterogeneity of Ca2+ release was severe when BCL was reduced to 300 msec (Fig. 2IIC), with highly variable release rates at the three randomly selected regions with slow or delayed release. There is clearly a high degree of variability in release rate all along the release front as indicated by the heterogeneity in fluorescence intensity immediately following the initiation of release (indicated by the white lines). This variability in release rate is further exaggerated during rapid pacing (Fig. 2IIC). These two figures are presented to show representative examples of 2‐D images and their accompanying linescans in order to demonstrate how the level of organization of the TT network affects SR Ca2+ release in Ca2+ transients. Similar results have been presented by other investigators in isolated myocytes of failing SHRs to those we found here in intact hearts (Song et al. 2006).