TABLE 1.
The effect of substrates and agents on changes in steady-state values of OCR, ISR, cytochrome c reduction, and NAD(P)H in rat pancreatic islets relative to the baseline (in the presence of 3 mm glucose)
Data were taken from Figs. 2–6, and each steady-state value shown was the average of the final 15 min of the indicated condition. Statistical analysis was carried out by comparing steady-state values before and after each change in medium composition using a paired t test. Abbreviations not defined in text: Ant A, antimycin A; Asc, ascorbate; Cyt c ox, cytochrome c oxidase; Glyc, glyceraldehyde.
| Process affected | Conditions | ΔOCR | ΔISR | ΔCytochrome c reduction | ΔNAD(P)H or Δcyt c ox reductiona |
|---|---|---|---|---|---|
| nmol/min/100 islets | ng/min/100 islets | % | |||
| Effect of increased Ca2+ influx in high glucose (Fig. 2A) | Condition 1: 20 mm Glc | +0.41 ± 0.032 (n = 4, p < 0.005) | +2.3 ± 0.35 (n = 6, p < 0.005) | +35 ± 4.3 (n = 4, p < 0.005) | NDb |
| Condition 2: 20 mm Glc +10 μm BayK 8644 | +0.074 ± 0.017 (n = 4, p < 0.05) | +3.1 ± 0.72 (n = 6, p < 0.01) | +5.2 ± 3 (n = 4) | ND | |
| Effect of increased Ca2+ influx in low glucose (Fig. 2B) | Condition 1: 3 mm Glc + 10 μm BayK 8644 | +0.025 ± 0.007 (n = 4, p < 0.05) | −0.02 ± 0.07 (n = 4) | −1.2 ± 0.31 (n = 3) | ND |
| Condition 2: 20 mm Glc + 10 μm BayK 8644 | +0.49 ± 0.038 (n = 4, p < 0.001) | +5.0 ± 1.0 (n = 4, p < 0.05) | +43 ± 1.9 (n = 3, p < 0.005) | ND | |
| Effect of increased TCA cycle activity in low glucose (Fig. 3A) | Conditon 1: 10 mm KIC | +0.33 ± 0.074 (n = 6, p < 0.005) | +2.3 ± 0.47 (n = 6, p < 0.005) | +28 ± 4.1 (n = 5, p < 0.005) | +25 ± 1.7 (n = 3, p < 0.005) |
| Condition 2: 10 mm KIC + 5 μm nimodipine | −0.097 ± 0.031 (n = 6, p < 0.005) | −2.5 ± 0.57 (n = 6, p < 0.01) | −6.4 ± 4.2 (n = 5) | −7.0 ± 1.0 (n = 3, p < 0.05) | |
| Effect of increased ETC in low glucose (Fig. 3B) | Cond 1: 10 mm Glyc | +0.13 ± 0.018 (n = 8, p < 0.005) | +0.9 ± 0.19 (n = 5, p < 0.005) | +15 ± 7.6 (n = 6, p < 0.05) | −7.0 ± 3.2 (n = 5) |
| Condition 2:10 mm Glyc + 5 μm nimodipine | −0.13 ± 0.011 (n = 8, p < 0.005) | −1.0 ± 0.17 (n = 5, p < 0.005) | −20 ± 7.2 (n = 6, p < 0.005) | +11 ± 4.8 (n = 5) | |
| Effect of reducing cytochrome c oxidase in low Glc (Fig. 4A) | Condition 1: 0.1 mm TMPD, 5 mm Asc | +0.40 ± 0.063c (n = 13, p < 0.005) | +0.39 ± 0.12 (n = 4) | +7.7 ± 7.0 (n = 4) | +94 ± 1.5a (n = 4, p < 0.0001) |
| Condition 2: 0.1 mm TMPD /5 mm Asc + 5 μm nimodipine | +0.094 ± 0.049c (n = 13) | +0.073 ± 0.031 (n = 4) | −4.7 ± 3.0 (n = 4) | −18 ± 12a (n = 4) | |
| Effect of reducing cytochrome c oxidase in high Glc (Fig. 4C) | Condition 1: 20 mm glucose | +0.26 ± 0.04 (n = 16, p < 0.0001) | +1.6 ± 0.1 (n = 5, p < 0.0001) | +37 ± 9.6 (n = 6, p < 0.01) | 7.5 ± 2.1 (n = 6, p < 0.01) |
| Condition 2: 20 mm Glc + TMPD/Asc (at 90 min) | +0.51 ± 0.06 (n = 16, p < 0.0001) | +0.96 ± 0.18 (n = 5, p < 0.01) | +4.0 ± 3.8 (n = 6) | 41.9 ± 8.7 (n = 6, p < 0.005) | |
| Effect of reducing cytochrome c with KCN (Fig. 5A, inset) | Condition 1: 20 mm Glc | +0.43 ± 0.016 (n = 6, p < 0.005) | +1.3 ± 0.13 (n = 6, p < 0.005) | +38 ± 6.4 (n = 6, p < 0.005) | ND |
| Condition 2: 20 mm Glc +1 μm KCN | −0.034 ± 0.009 (n = 6, p < 0.05) | +0.70 ± 0.24 (n = 6, p < 0.05) | +6.9 ± 1.0 (n = 6, p < 0.005) | ND | |
| Effect of oxidizing cytochrome c with antimycin A (Fig. 5A, inset) | Condition 1: 20 mm Glc | +0.50 ± 0.062 (n = 3, p < 0.05) | +2.2 ± 0.42 (n = 3, p < 0.05) | +41 ± 2.6 (n = 3, p < 0.005) | ND |
| Condition 2: 20 mm Glc + 0.6 ng/ml Ant A | −0.024 ± 0.002 (n = 3, p < 0.01) | +0.13 ± 0.2 (n = 3) | −1.4 ± 1.8 (n = 3) | ND | |
| Effect of glucose stimulation and no further additions (Fig. 5A) | Condition 1: 20 mm glucose | +0.36 ± 0.02 (n = 12, p < 0.0001) | +2.0 ± 0.2 (n = 6, p < 0.0005) | +42 ± 2.5 (n = 12, p < 0.0001) | ND |
| Condition 2: 20 mm Glc (at time = 90) | −0.006 ± 0.007 (n = 12) | +0.31 ± 0.23 (n = 6) | +2.1 ± 1.4 (n = 12) | ND | |
| Effect of reducing cytochrome c with KCl in high Glc/diazoxide (Fig. 6A) | Condition 1: 20 mm Glc | +0.28 ± 0.024 (n = 8, p < 0.005) | +0.25 ± 0.075 (n = 5, p < 0.05) | +28 ± 3.9 (n = 6, p < 0.005) | ND |
| Condition 2: 30 mm KCl + 20 mm Glc | +0.12 ± 0.014 (n = 8, p < 0.005) | +4.2 ± 0.66 (n = 5, p < 0.01) | +15 ± 3.2 (n = 6, p < 0.05) | ND | |
| Effect of increasing cytochrome c reduction with KCl at low Glc/diazoxide (Fig. 6D) | Cond 1: 30 mm KCl + 3 mm glucose | +0.051 ± 0.004 (n = 6, p < 0.0001) | +0.48 ± 0.14 (n = 4, p < 0.05) | +8.7 ± 4.4 (n = 6, p < 0.05) | ND |
| Condition 2: 20 mm Glc + 30 mm KCl | +0.27 ± 0.018 (n = 6, p < 0.0001) | +1.9 ± 0.4 (n = 4, p < 0.05) | +24.8 ± 9.0 (n = 6, p < 0.005) | ND | |
a Cytochrome c oxidase reduction was measured and not NAD(P)H.
b ND, not determined.
c OCR measured by the Seahorse was normalized by dividing by the OCR obtained at 3 mm glucose and is unitless.