Blunting of rapid onset vasodilatation and blood flow restriction in arterioles of exercising skeletal muscle with ageing in male mice

Abstract

Exercise capacity and skeletal muscle blood flow are diminished with ageing but little is known of underlying changes in microvascular haemodynamics. Further, it is not clear how the sympathetic nervous system affects the microcirculation of skeletal muscle with ageing or whether sex differences prevail in the regulation of arteriolar diameter in response to muscle contractions. In the gluteus maximus muscle of C57BL/6 mice, we tested the hypothesis that ageing would impair 'rapid onset vasodilatation' (ROV) in distributing arterioles (second-order, 2A) of old (20-month) males (OM) and females (OF) relative to young (3-month) males (YM) and females (YF). Neither resting (approximately 17 microm) nor maximum (approximately 30 microm) 2A diameters differed between groups. In response to single tetanic contractions at 100 Hz (duration, 100-1000 ms), ROV responses were blunted by half in OM relative to OF, YM or YF. With no effect in YM, blockade of alpha-adrenoreceptors with phentolamine (1 mum) restored ROV in OM. Topical noradrenaline (1 nM) blunted ROV in YM and YF to levels seen in OM and further suppressed ROV in OM (P < 0.05). To evaluate arteriolar blood flow, red blood cell velocity was measured in 2A of OM and YM; respective heart rates (353 +/- 22 vs. 378 +/- 15 beats min(1)) and carotid arterial blood pressures (76 +/- 3 vs. 76 +/- 1 mmHg) were not different. Blood flows at rest (0.6 +/- 0.1 vs. 1.6 +/- 0.2 nl s(1)) and during maximum dilatation (2.0 +/- 0.8 vs. 5.4 +/- 0.8 nl s(1)) with sodium nitroprusside (10 microM) were attenuated >60% (P < 0.05) in OM. Blood flow at peak ROV was blunted by 75-80% in OM vs. YM (P < 0.05). In response to 30 s of rhythmic contractions at 2, 4 and 8 Hz, progressive dilatations did not differ with age or sex. Nevertheless, resting and peak blood flows in YM were 2- to 3-fold greater (P < 0.05) than OM. We suggest that ageing blunts ROV and restricts blood flow to skeletal muscle of OM through subtle activation of alpha-adrenoreceptors in microvascular resistance networks.

Figure 1. Rapid onset vasodilatation in response to brief tetanic contraction

Representative traces of diameter responses to single tetanic contractions of 100 ms (left) and 1000 ms (right) duration from a young male mouse. Vertical bars indicate stimulation at 100 Hz. As ROV was nearly instantaneous, delays in diameter responses reflect time to refocus the microscope after contraction.

Figure 2. Blunting of ROV in old male mice

A, across the range of contraction durations, 2A diameter change in response to single tetanic contractions was attenuated by nearly half in OM as compared to YM, YF or OF, (n= 6–18 per group). *P < 0.05, OM vs. YM and YF; #P < 0.05, OM vs. other groups. B, in each experimental group, the duration of muscle contraction correlated well with the change in arteriolar diameter (n= 6–18 per group). However the slope of ROV responses to increasing stimulus duration was blunted in OM. *P < 0.05, OM vs. other groups. Abbreviations: YM, young male; YF: young female; OM: old male; OF: old female.

Figure 3. Roles of α-adrenoreceptor inhibition and activation in ROV

A, superfusion of phentolamine (1 μm) increased ROV in OM to levels not different from those recorded in YM. Conversely, superfusion of NA (1 nm) blunted ROV in YM to levels not different from those recorded in OM. Resting diameters (Table 1) were not different between conditions. *P < 0.05 for OM and YM + NA vs. YM and OM + phentolamine (n= 6–8 per group). B, superfusion of NA (1 nm) blunted ROV in YF (consistent with its effect in YM, panel A), particularly at longer durations of contraction. However, superfusion of phentolamine (1 μm) had no effect on ROV responses in OF, which were not different from those in YF. Resting diameters (Table 1) were not different between conditions. *P < 0.05, YF + NA vs. other groups (n= 4–6 per group). Abbreviations are defined in Fig. 2 legend.

Figure 4. Effects of NA and phentolamine on ROV in Young and Old male mice

A, young males: superfusion of NA (1 nm) blunted ROV (*P < 0.01) while superfusion of phentolamine (1 μm) had no significant effect on ROV (n= 5). B, old males: superfusion of NA (1 nm) blunted ROV (*P < 0.01) while superfusion of phentolamine (1 μm) enhanced ROV vs. OM under control conditions (#P < 0.01; n= 5). C, comparisons of YM vs. OM (data in A and B re-plotted): under control conditions, ROV was blunted in OM vs. YM (*P < 0.01). For YM + NA, ROV was blunted significantly below control (†P < 0.01) and these levels were not significantly different from OM. For OM + NA, ROV was further depressed for each contraction (#P < 0.01). Phentolamine had no effect in YM yet increased ROV significantly in OM (‡P < 0.01). Note that 1 OM had a particularly large response to 1000 ms contraction with phentolamine that elevated the mean ±s.e. for this data point.

Figure 5. Arteriolar haemodynamic responses during ROV were blunted in OM vs. YM

A, at rest and during peak dilatation in response to single tetanic contractions, blood flow was greatly attenuated in OM vs. YM. *P < 0.05, OM vs. YM. B, mean red blood cell velocity during peak dilatation did not change with increasing stimulus duration in either YM or OM. However, baseline and response values were blunted consistently in OM vs. YM. *P < 0.05, OM vs. YM. C, wall shear rate during peak dilatation did not change with increasing stimulus duration in either YM or OM. However, WSR was consistently lower in OM vs. YM. *P < 0.05, OM vs. YM. For all panels, n= 6 per group. Abbreviations are defined in Fig. 2 legend.

Figure 6. With rhythmic contractions, arteriolar dilatations are independent of age or sex yet blood flow is blunted in OM

A, with no difference in resting or maximal diameters (Table 1), the magnitude of arteriolar dilatation increased with contraction frequency in a similar manner for each experimental group. *Main effect of stimulus frequency, P < 0.05 (n= 6–18 per group). B, blood flows at rest (baseline) and in response to 30 s of rhythmic contractions were restricted by more than half in OM compared to YM. *P < 0.05, OM vs. YM (n= 6 per group). Abbreviations are defined in Fig. 2 legend.