Vascular Smooth Muscle Glycogen Metabolism Studied by ¹³C-NMR

Abstract

Vascular smooth muscle glycogen stores are traditionally thought to be small compared to other glycogen-containing tissues such as striated muscle or liver. However, glycogen has been thought to be an important carbon substrate for oxidative metabolism in support of contraction in vascular smooth muscle. We examined the synthesis and degradation of glycogen in isometrically mounted hog carotid artery using ¹³C-NMR spectroscopy. The rate of net glycogen synthesis from 1-¹³C-glucose was found to be constant during the first 8 h of incubation of carotid arteries with 10 mM glucose at 37 °C and then decreased towards a rate of zero by 14 h of incubation. During 8 h of incubation in the presence of 5 mM glucose, the content of glycogen increased from 1.5 to 8.1 µmol/g blot weight in the absence of insulin and to 11.4 µmol/g blot weight in the presence of 0.5 U/ml insulin. During prolonged glycogen loading, there was a simultaneous degradation of previously synthesized 6-¹³C-glycogen during synthesis of 1-¹³C-glycogen from 1-¹³C-glucose indicating substrate cycling of glycogen metabolism. This substrate cycling results in a pattern of glycogen utilization in which the most recently synthesized glucosyl units of glycogen are utilized only slightly more readily than the previously synthesized glucosyl units of glycogen. We conclude that glycogen stores are larger and more dynamic than previously thought in vascular smooth muscle consistent with an important role for glycogen as a carbon source for smooth muscle energy metabolism.

Oleate Oxidation and Mitochondrial Substrate Selection in Vascular Smooth Muscle

“…It has been shown that VSM has the ability to synthesize glycogen at a rate of 0.93 µmol glucosyl units of glycogen/gram wet weight (w.w.)/hour for the first 6 h of the glycogen synthesis incubation in PSS containing 5 m M glucose [14]. For every experiment, one section (30–70 mg) from each artery was removed prior to experimental manipulation for glycogen synthesis incubation.…”

Influence of glycogen storage on vascular smooth muscle metabolism

“…Hog carotid arteries were then incubated in PSS containing 5 mM glucose for 12-16 h at 37°C before isometric contraction, which resulted in glycogen concentrations ranging from 7 to 15 µmol glucosyl units glycogen/g wet wt of tissue (µmol/g). This range was selected on the basis of the observation that hog carotid artery can synthesize at least 0.93 µmol glycosyl units of glycogen • g Ϫ1 •h Ϫ1 in the first 6 h of incubation under these conditions (13). Therefore, arteries that did not synthesize at least 6 µmol/g of new glycogen were considered nonviable and were excluded, and only tissues with glycogen contents above 7 µmol/g (1 µmol/g limit dextrin and 6 µmol/g newly synthesized glycogen) were used in further analysis.…”

“…After 6 h, three sections from each artery (ϳ30 mg each) were removed, blotted dry, weighed, and frozen in liquid nitrogen for subsequent analysis of glycogen content (final glycogen content). The remaining portion of the artery (ϳ200 mg) was blotted dry, weighed, and frozen in liquid nitrogen for subsequent extraction and 13 C nuclear magnetic resonance (NMR) analysis. Aliquots (1.5 and 4.0 ml) of the superfusate for each tissue were collected both before and after isometric contraction to assay for [1-13 C]glucose flux, total lactate production, and 1 H NMR spectroscopy.…”

“…An additional three sections from each artery (ϳ30 mg each) were placed in separate incubation chambers in PSS containing 5 mM glucose for 6 h. Tissues that did not synthesize at least 6 µmol/g glucosyl units of glycogen during this incubation were excluded from further analysis. This was done to ensure that tissues used for subsequent 13 C NMR analysis were viable and therefore appropriate representatives for measurement of substrate metabolism of VSM with low levels of glycogen.…”

“…This rapid depletion of glycogen substrate reserves raises speculation regarding the role of glycogen as an important oxidative substrate for VSM. However, it has been shown that VSM glycogen stores can accumulate approximately fivefold higher glycogen contents on incubation in 5 mM glucose for 12-16 h at 37°C (13). In addition, it has been shown that the glycogen utilization is directly proportional to the tissue glycogen content (15), suggesting that glycogen may play a much larger role in VSM metabolism than previously thought.…”