| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
,2
* Dipartimento di Scienze della Vita, Seconda Università degli Studi di Napoli, Caserta, Italy;
Dipartimento di Scienze Biologiche ed Ambientali, Università degli Studi del Sannio, Benevento, Italy; and
Dipartimento delle Scienze Biologiche, Sez. Fisiologia ed Igiene, Università degli Studi di Napoli "Federico II," Napoli, Italy
2Correspondence: A.L. Dipartimento di Scienze della Vita, Seconda Università degli Studi di Napoli, Via Vivaldi 43, 81100, Caserta, Italy; E-mail: antonia.lanni{at}unina2.it, or F.G. Dipartimento di Scienze Biologiche ed Ambientali, Università degli Studi del Sannio, Via Port’Arsa 11, 82100, Benevento, Italy; E-mail: goglia{at}unisannio.it
SPECIFIC AIMS
The metabolic response to food deprivation involves hormonal and metabolic adaptations accompanied by transcriptional reprogramming events. These adaptations can be divided into three phases: the first is a brief period of adaptation marked by glycogen deprivation, the second involves protein sparing with an associated use of lipids as the main fuel, and the third is characterized by a metabolic and endocrine shift, resulting in an increase in protein utilization. The aim of the present study was to shed light on the temporal sequence of biochemical and molecular events (and their interrelationships) leading to adaptive responses at the morphological/functional level in gastrocnemius muscle.
PRINCIPAL FINDINGS
1. Food deprivation rapidly induces reprogramming of the rat’s gastrocnemius muscle toward increased lipid metabolism
Changes in gastrocnemius muscle toward increased use of lipids as fuel begin with a rapid recruitment of factors that trigger this reprogramming. Nuclear accumulation within 6 h was observed of peroxisome proliferator-activated receptor (PPAR) 
coactivator (PGC)-1
and PPAR 
proteins (nuclear protein levels increased by 4- and 2-fold, respectively). This accumulation was transient, as cytosolic and nuclear protein levels declined rapidly to control levels at 48 h. The increase in nuclear levels of PGC-1 and PPAR coincided with an increase in mRNA levels for the transcription factor MyoD, also involved in muscular reprogramming and an essential factor for the transcription of uncoupling protein UCP3, and of genes involved in mitochondrial lipid handling, including UCP3, carnitine palmitoyl transferase (CPT) 1b, mitochondrial thioesterase (MTE) I (Fig. 1
A). This was followed by an increase at 12 h in mitochondrial protein levels (UCP3 by 5-fold, MTE I by 3-fold, Fig. 1B
) with a concomitant increase in the rate of mitochondrial fatty acid oxidation (Fig. 1C
).
|
2. Upon food deprivation, muscle fiber composition rapidly starts to be reprogrammed at the molecular level toward type I oxidative fibers
The increased nuclear accumulation of PGC-1 and PPAR proteins coincided with a sharp increase in myosin heavy chain (MHC) Ib mRNA and protein (by 8-fold) within 6 h. During the fasting period, MHC Ib levels remained elevated, leading to an increase in type I fiber content at 48 h as well as a redder appearance of the whole muscle, with a reduced size of the white sections.
3. Recruitment of AMP-activated protein kinase (AMPK) in muscular cell signaling on food deprivation
Within 6 h, muscle AMP levels rose by 2-fold. Figure 2
shows that, in concomitance, phosphorylation of AMPK increased by 4-fold within 6 h whereas total AMPK protein levels remained unaltered. AMPK phosphorylation was consistent with an increased phosphorylation (and inactivation) of acetyl coenzyme A (CoA) carboxylase (ACC), thus inhibiting the formation of malonyl CoA and inducing increased activity of the CPT system, and consequently of mitochondrial fatty acid uptake and increased oxidation, starting from 12 h. Phosphorylation of AMPK was transient, the increase above control levels being 
2-fold at 12 h and zero at 24 h. These findings provide clear evidence for a rapid involvement of AMPK in skeletal muscle’s usage of fat as fuel after a fasting stimulus.
|
4. Fatty acid levels and AMPK phosphorylation are functionally linked
Serum fatty acid levels were elevated at 12 h, peaked at 24 h, and had declined to 2.5-fold that of the fed control level at 48 h (Fig. 2B
). The increase in serum fatty acid levels coincided with a decrease in gastrocnemius muscle AMPK phosphorylation. Since AMPK is known for its role as fuel provider, it seems apparent that once fatty acid levels are increased, kinase activity is inhibited. The results indicate that an increase in serum fatty acid levels may be the cause of the inhibition of AMPK activity. Such a functional-linked phenomenon is demonstrated when removal of fatty acids from the serum by treatment with nicotinic acid (NA) restores muscular AMPK phosphorylation. These data demonstrate an active role for fatty acids in modulating AMPK activity, and thus in mitochondrial fatty acid uptake.
CONCLUSIONS AND SIGNIFICANCE
Recent evidence suggests that muscle structure and metabolic activity are tightly linked at the molecular level. The nuclear receptor PPAR plays a pivotal role in the control of the mitochondrial oxidation of fatty acids, the most important metabolic process by which fatty acids are used to provide energy for the heart and skeletal muscles. In food deprivation, how well an organism copes or even whether it survives depends on its capacity to make appropriate metabolic adjustments, the ability to oxidize fatty acids being crucial. The data of this study illustrate the capacity of rat gastrocnemius muscle to be rapidly reprogrammed at the metabolic level upon food deprivation and highlight roles played by PGC-1, PPAR , and MyoD as in vivo determinants of such reprogramming. Indeed, their increased expression and nuclear accumulation immediately after the food deprivation stimulus are followed by increased mRNA levels for genes involved in mitochondrial lipid metabolism and fatty acid oxidation (CPT, UCP3, and MTE I) and rapid increases in the levels of the mRNA and protein for the MHCIb gene, indicative of the formation of oxidative fibers. In contrast to data obtained earlier from mice, we report here that in the rat in vivo a rapid, transient phosphorylation of AMPK occurs, consistent with a rise in AMP levels, highlighting a role for AMPK in the rat gastrocnemius muscle response to food deprivation. In concert with this AMPK phosphorylation, there is increased phosphorylation of ACC, a downstream target of AMPK. These data, showing a coordinated activation of AMPK and ACC in the metabolic pathway leading to increased mitochondrial fatty acid oxidation, point to AMPK participating in nutritional energy-sensing in vivo as it does in vitro. In addition, AMPK phosphorylation showed a negative correlation with the increase in the circulating fatty acid level. At the time this level (Fig. 2B
) mitochondrial fatty acid oxidation (Fig. 1C
) began to increase, and AMPK phosphorylation started to decrease (Fig. 2A
). This process was reversed by NA treatment, demonstrating that fatty acids are most likely involved in AMPK activation/deactivation.
The findings reported here elucidate the physiological sequence of events in the in vivo coordinated pattern of biochemical/molecular changes involved in the reprogramming of gastrocnemius muscle after food deprivation, highlighting the roles played in vivo by PPAR , AMPK, and fatty acids. Our results also provide insight into the in vivo modulation of skeletal muscle AMPK activity in situations involving energy shortage in a physiological model.
|
FOOTNOTES
1 These authors contributed equally to this work 
To read the full text of this article, go to http://www.fasebj.org/cgi/doi/10.1096/fj.06-6025fje
This article has been cited by other articles:
|
|
 |
|
  P. de Lange, R. Senese, F. Cioffi, M. Moreno, A. Lombardi, E. Silvestri, F. Goglia, and A. Lanni Rapid Activation by 3,5,3'-L-Triiodothyronine of Adenosine 5'-Monophosphate-Activated Protein Kinase/Acetyl-Coenzyme A Carboxylase and Akt/Protein Kinase B Signaling Pathways: Relation to Changes in Fuel Metabolism and Myosin Heavy-Chain Protein Content in Rat Gastrocnemius Muscle in Vivo Endocrinology, December 1, 2008; 149(12): 6462 - 6470. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 Pieter de Lange, M. Moreno, E. Silvestri, A. Lombardi, F. Goglia, and A. Lanni Fuel economy in food-deprived skeletal muscle: signaling pathways and regulatory mechanisms FASEB J, November 1, 2007; 21(13): 3431 - 3441. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
