Study reveals molecular mechanisms behind hibernation in mammals

Researchers have shed light on the molecular mechanisms underlying hibernation, publishing their findings today as a revised preprint in eLife.

Their research, on small and large hibernating mammals, is described by the editors as an important study advancing our knowledge of the role of myosin structure and energy consumption on the molecular mechanisms of hibernation, supported through solid methodology and evidence. The results also suggest that myosin, a type of motor protein involved in muscle contraction, plays a role in non-shivering thermogenesis during hibernation, where heat is produced independently of shivering-related muscle activity.

Hibernation is a survival strategy used by many animals, characterized by a state of deep dormancy and profound reductions in metabolic activity, body temperature, heart rate, and respiration. During hibernation, animals rely on stored energy reserves, particularly fat, to maintain bodily functions. Metabolic slowing allows hibernators to conserve energy and endure long periods of food shortage and harsh environmental conditions during winter. However, the cellular and molecular mechanisms underlying hibernation remain incompletely understood.

Hibernating small mammals experience prolonged episodes of a hypometabolic state called torpor, which significantly decreases their body temperature and is punctuated by spontaneous periods of interepisode euthermic arousals (IBA) – during which they temporarily increase their body temperature to restore certain physiological functions, such as eliminating waste and eating more food.

This contrasts with larger mammals, whose body temperature is reduced much less during hibernation and remains fairly constant. Skeletal muscle, which makes up about half of a mammal’s body mass, plays a key role in determining its heat production and energy consumption.

“Until recently, it was thought that the energy consumption of skeletal muscles was mainly linked to the activity of myosin, involved in muscle contraction. However, there is growing evidence that even when they are relaxed “, skeletal muscles still use a small amount of energy,” says lead author Christopher Lewis, a postdoctoral researcher in the Department of Biomedical Sciences at the University of Copenhagen, Denmark.

“Myosin heads in passive muscles can be in different resting states: the ‘disordered-relaxed’ state or DRX, and the ‘super relaxed’ state or SRX. Myosin heads in the DRX state use ATP, the cell’s energy currency, between five and ten times faster than those in the SRX state,” adds Lewis.

Lewis and colleagues hypothesized that changes in the proportion of myosin in DRX or SRX states could contribute to the reduction in energy consumption observed during hibernation. To test this, they took skeletal muscle samples from two small hibernators – the thirteen-lined ground squirrel and the garden dormouse – and two large hibernators – the American black bear and the brown bear.

First, they sought to determine whether myosin states and their respective ATP consumption rates were different between periods of activity and hibernation. They examined muscle fibers taken from the two bear species during their summer active (SA) phase and their winter hibernation period.

They found no difference in the proportion of myosin in the DRX or SRX state between the two phases. To measure the rate of ATP consumption by myosin, they used a specialized test called the Mant-ATP flush test. This revealed that there was also no change in myosin energy consumption rates. This could help prevent the occurrence of significant muscle wasting in bears during hibernation.

The team also conducted the Mant-ATP chase assay on samples collected from the small mammals during AS, IBA, and torpor. As in larger hibernators, they observed no difference in the percentage of myosin heads in the SRX or DRX formation between the three phases. However, they found that the ATP turnover time of myosin molecules in both formations was lower in IBA phase and torpor compared to SA phase, leading to an unexpected overall increase in ATP consumption .

Because small mammals experience a greater drop in body temperature during hibernation than larger mammals, the team tested whether this unexpected increase in ATP consumption also occurred at a lower temperature. They re-ran the Mant-ATP chase assay at 8°C, compared to the 20°C laboratory room temperature used previously. Lowering temperature reduced the turnover times of DRX- and SRX-bound ATP in SA and IBA, leading to increased ATP consumption.

It is well known that metabolic organs, such as skeletal muscles, increase core body temperature in response to significant cold exposure, either by inducing shivering or by non-shivering thermogenesis. Cold exposure caused an increase in ATP consumption by myosin in samples obtained during AS and IBA, suggesting that myosin may contribute to non-shivering thermogenesis in pups. hibernators.

The team did not observe cold-induced changes in myosin energy consumption in samples obtained during torpor. They suggest that this is likely a protective mechanism aimed at keeping body temperature low and broader metabolic shutdown observed during torpor.

Finally, the researchers wanted to understand the changes that occur at the protein level during the different phases of hibernation. They assessed whether hibernation affects the structure of two thirteen-lined ground squirrel myosin proteins, Myh7 and Myh2. Although they observed no hibernation-related changes in the structure of Myh7, they found that Myh2 underwent significant phosphorylation – a process crucial for energy storage – during torpor, compared to SA and IBA.

They also analyzed the structure of the two proteins in brown bears, finding no structural differences between AS and hibernation. They therefore conclude that Myh2 hyperphosphorylation is specifically associated with torpor, rather than hibernation in general, and propose that this serves to increase myosin stability in small mammals. This could act as a potential molecular mechanism to attenuate myosin-associated increases in skeletal muscle expenditure in response to cold exposure during periods of torpor.

eLifeThe editors note that some areas of the study merit further study. Namely, the muscle samples were taken exclusively from the legs of the animals studied. Since the body and limbs have different temperatures, studying muscle samples from other areas of the body would further validate the team’s findings.

“In total, our results suggest that adaptations of ATP turnover in DRX and SRX myosin states occur in small mammals like the thirteen-lined ground squirrel during hibernation in cold environments. In contrast, mammals “Largers like the American black bear show no such changes, probably due to their stable body temperature during hibernation,” concludes lead author Julien Ochala, associate professor in the Department of Biomedical Sciences at the University of Copenhagen. “Our results also suggest that myosin may contribute to non-shivering thermogenesis of skeletal muscles during hibernation.”