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Novel technologies and measurement of microvascular blood flow in muscle


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Clark, ADH 2005 , 'Novel technologies and measurement of microvascular blood flow in muscle', PhD thesis, University of Tasmania.

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In the field of microvascular blood flow, particularly to the skeletal muscle, there has
been a considerable history of reference to the incongruity of total limb blood flow and
actual myocyte perfusion. Embedded in such an idea is the concept that flow to skeletal
muscle should be composed of two physiologically distinct circulations — one that
perfuses the myoctes (and therefore may be called "nutritive") and another which must
act as a form of physiological shunt or "non-nutritive". The "nutritive and non-nutritive
flow" hypothesis was renewed by the muscle research group of the Biochemistry
Department at the University of Tasmania to explain perfusion dependent metabolic
effects of vasoconstrictors on skeletal muscle.
This thesis focuses on the assessment of microvascular perfusion, compared to total flow,
in skeletal muscle and in particular in relation to reports of direct and indirect effects of
insulin to increase microvascular perfusion of this tissue in vivo. Initially laser Doppler
Fluxmetry technology (LDF) was applied to estimate muscle perfusion. For the purposes
of verification and comparison two other techniques under development by our group,
contrast enhanced ultrasound (CEU) and 1-methyl xanthine metabolism, were also used.
In the first section of this thesis the variable of total flow is eliminated by using a constant
flow (pump perfused) rat hindlimb. During the application of vasoconstrictors that
caused either predominantly nutritive or non-nutritive flow LDF estimation of perfusion
was found to be highly variable in spite of the constant total flow. LDF comprises a non
vectorial voltage measurement generated by particle movement through a volume of
tissue illuminated by the laser light. LDF measurement using a large probe was highly correlated with the metabolic changes
induced by changes in nutritive and non-nutritive flow. However, LDF measurement
using a smaller implantable probe which measures particle movement in a smaller
volume of muscle tissue, showed that there is considerable heterogeneity of response.
To better understand the factors that influence changes in the LDF measurement LDF
measurements were made of flow controlled through fabricated glass capillary arrays of
various architectures.
The third study addressed changes induced by insulin in vivo, in anaethetised rats during
a hyperinsulinaemic euglycaemic clamp. Using a large surface LDF probe the LDF
signal from the muscle increased during the administration of insulin. In contrast the
LDF signal did not increase during epinephrine administration even though the
epinephrine caused a comparable increase in femoral artery blood flow as seen with
insulin. Furthermore the time course of LDF change in the presence of insulin was more
closely aligned with changes in glucose uptake than it was with changes in femoral blood
In the final series of experiments using LDF, changes due to insulin in human muscle was
assessed. The results suggested that physiological hyperinsulinaemia stimulated not only
total blood flow and skin microvascular perfusion, but also augmented human skeletal
muscle microvascular recruitment and vasomotion as detected directly by laser Doppler
In the second technique, CEU, microbubbles (an albumin membrane encapsulating an
inert gas in the form of a 4 micron diameter sphere) act as contrast when tissue is
visualised using ultrasonography. Analysis of the rate and extent of contrast
(microbubble) appearance during imaging of the target tissue allows us to calculate
indicators of the microvascular velocity and microvascular volume.
The third technique based on capillary endothelial metabolism of the exogenous substrate
1-methylxanthine, was developed at the University of Tasmania. 1-Methylxanthine is a
reporter substrate, which as it passes through the nutritive capillary route of the muscle is
metabolized to 1-methyl urate; metabolism is proportional to available capillary surface
area. To consolidate the LDF findings, the two additional techniques for assessing changes in
capillary recruitment were used. Once again, the hyperinsulinaemic euglycaemic clamp
was used but on this occasion a physiologic dose of insulin was infused. Microvascular
recruitment (measured by 1-MX metabolism as well as by CEU) increased during insulin administration. Furthermore, at certain time points microvascular recruitment occurred
without changes to femoral artery flow. From these studies it was evident that insulin
increases tissue perfusion by recruiting microvascular beds, and at physiological
concentrations this precedes increases in total muscle blood flow by 60-90 min. Since
there was no change in the mean red cell velocity, it seem likely that insulin had
increased capillary recruitment by redistributing blood flow from other vessels possibly
In the final study capillary recruitment was again measured by the metabolism of infused
1-methyl xanthine and used to assess responses to insulin in normal colony rats as well as
lean and obese Zucker rats (an animal model for type 2 diabetes). It was concluded that
muscle insulin resistance of obese Zucker rats is accompanied by impaired hemodynamic
responses to insulin including markedly impaired capillary recruitment and femoral blood
flow when compared to lean Zuckers or colony control rats.
Taken together, the findings embodied by this thesis show that the three techniques of
LDF, CEU and 1-MX metabolism, each detect a vascular effect of insulin in vivo,
characterized by capillary recruitment. This was detectable at physiologic insulin and
occurred in both muscle of rats and humans. In addition, LDF added a further aspect of
insulin action of an insulin-mediated increase in vasomotion at low frequency, very likely
suggesting increased neural activity.

Item Type: Thesis - PhD
Authors/Creators:Clark, ADH
Keywords: Laser Doppler blood flowmetry, Microcirculation, Oxygen, Muscles, Blood flow
Copyright Holders: The Author
Copyright Information:

Copyright 2005 the Author - The University is continuing to endeavour to trace the copyright
owner(s) and in the meantime this item has been reproduced here in good faith. We
would be pleased to hear from the copyright owner(s).

Additional Information:

Thesis (Ph.D.)--University of Tasmania, 2005. Includes bibliographical references

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