Ground squirrels spend late summer gorging on food, preparing for hibernation. They need to store a lot of energy in the form of fat, which becomes their main source of fuel underground in their hibernation burrows throughout the winter.
During hibernation, ground squirrels enter a state called torpor. Their metabolism drops to just 1% of summer levels and their body temperature can dive to near zero. Torpor greatly reduces the amount of energy the animal needs to stay alive until spring.
This long fast has a downside: no new intake of protein, which is crucial for maintaining body tissues and organs. This is a particular problem for the muscles. In humans, long periods of inactivity, such as prolonged bed rest, lead to muscle wasting. But muscle wasting is minimal in hibernating animals. Despite six to nine months of inactivity and no protein intake, they preserve muscle mass and performance remarkably well – a very practical adaptation that helps ensure a successful spring breeding season.
How do hibernators do? It was a real headache for hibernation biologists for decades. Our research tackle team this question by investigating how hibernating animals might get major help microbes that live in their guts.
A nitrogen recycling system
We knew from previous research that a hibernating the gastrointestinal system undergoes dramatic changes in its structure and function, from summer feeding to winter fasting. And it’s not just animals that fast all winter, their gut microbes do too. With our collaborators in microbiology, we discovered that Winter fasting alters the gut microbiota a little.
And then we wondered…could gut microbes play a functional role in the hibernation process itself? Could certain bacteria help keep muscles and other tissues functioning when the mostly immobile animals aren’t eating?
Biologists had previously identified a clever trick in ruminants, such as cattle, that helps them survive when protein intake in the diet is low or protein requirements are particularly high, such as during pregnancy. A process called urea nitrogen recovery allows the animal to scavenge nitrogen – an essential ingredient for building protein – that would otherwise be excreted in the urine as waste urea. Instead, urea nitrogen is retained in the body and used to make amino acids, the building blocks of protein.
This recovery operation depends on the chemical breakdown of urea molecules to release their nitrogen. But here’s the catch: the chemical breakdown of urea requires urease, an enzyme that animals don’t produce. So how does a cow, for example, extract this nitrogen from urea?
It turns out that certain microbes that are normal residents of animal intestines can do just that. They make the enzyme urease and use it to chemically separate urea molecules, releasing nitrogen, which becomes part of the ammonia molecules. The microbes then absorb the ammonia and use it to make new proteins themselves.
The peculiarities of the digestive system of ruminants allow these animals to benefit greatly from this process. But for other animals – like hibernators and us – it was less clear if and how urea nitrogen could enter animals’ bodies to support protein synthesis.
This was our challenge as scientists: could we demonstrate the recycling of urea nitrogen in hibernators and show that it is particularly useful for them to be longer to fast?
Our experimental game plan
Using the 13-line ground squirrel, we designed experiments to study key steps in urea nitrogen recovery.
First, we injected urea molecules into the squirrel’s blood in which the two nitrogen atoms were replaced by a heavier form of nitrogen that is only naturally present in small amounts. in the body.
We were able to track these heavier nitrogen atoms as the injected urea moved from the blood into the gut, then as microbial urease broke down the urea into its component parts, and finally into metabolites and proteins in the squirrels’ tissues. Wherever we saw higher levels of the heaviest form of nitrogen, we knew that urea was the source of the nitrogen, and therefore gut microbes must be responsible for bringing urea nitrogen back into the animal bodies.
To confirm that microbes were performing nitrogen recycling, we compared squirrels that had normal gut microbiomes to squirrels that did not. We treated some animals with antibiotics to reduce gut microbes at three times of the year: summer; at the beginning of winter, when they were one month away from fasting and hibernation; and the end of winter, when they were four months away from fasting and hibernation.
In squirrels with normal microbiomes, we saw evidence of urea nitrogen scavenging at every step of the process we tested. But squirrels with depleted microbiomes showed minimal urea nitrogen recovery. Our observations confirmed that this process did depend on the ability of gut microbes to break down urea and release its nitrogen in the hibernating intestines. The liver and muscle tissues of the hibernators incorporated the most urea nitrogen at the end of winter, that is, the longer they had hibernated and without food.
We also found that ground squirrels contribute to this remarkable symbiosis. During hibernation, their gut cells increase the production of proteins called urea transporters. These molecules are housed in the intestinal cell membranes and transport urea from the blood to the intestine where the microbes that contain urease are found. This aid means that the little urea the animal produces during hibernation has an easier path to the intestine.
Finally, we discovered that it wasn’t just squirrels that benefited from this process. The microbes also used urea nitrogen to build their own proteins, showing that scavenging urea nitrogen provides both parties with this important molecular building block during the long winter fast.
Could this kind of symbiosis help humans?
This example of hibernator-microbe symbiosis has potential clinical applications. For example, undernourishment, which affects millions of people around the world, leads to a progressive loss of muscle mass and compromises health. Sarcopenia, which is muscle wasting that is a natural part of aging, impairs mobility and makes people more susceptible to injury. A detailed understanding of how the hibernator nitrogen scavenging system is most effective when the risk of tissue loss and muscle wasting is highest could lead to new therapies to help people in situations similar.
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Another potential application is in manned spaceflight, where crew members experience high rates of muscle atrophy due to a microgravity-induced suppression of muscle protein synthesis. Even the extensive exercise regimen that astronauts undertake to compensate for this is insufficient. A microbiome-based countermeasure that facilitates muscle protein synthesis similar to the process we observed in hibernators might be worth investigating.
These applications, although theoretically possible, are still far from being delivered. But studies in the 1990s showed that humans are capable of recycle small amounts of urea nitrogen using their gut microbes. The necessary machinery is therefore in place; all that remains is to optimize it.
