Nuclear Magnetic Resonance

A view inside – A fish in the NMR

What goes through a fish’s head when it swims through acidic water? This question has been occupying the biologist Matthias Schmidt for some time. As part of his doctorate at the Alfred Wegener Institute he is investigating how the blood flow of a fish alters as the carbon dioxide concentration increases in the water. With the assistance of magnetic resonance imaging he is able to peer into the workings of a fish.

A red and white triangle with a large exclamation mark hangs on the heavy safety door to the MRT laboratory on the ground floor of the Alfred Wegener Institute, warning against strong magnetic fields and danger! A second sign warns that metallic objects near the magnets make for dangerous projectiles. People with pacemakers and other metallic implants may not even enter the laboratory – all others must wait until someone behind the closed door hears the knocking.

Matthias Schmidt opens the door to the large laboratory room only a little later. In his white coat, the young biologist looks like an X-ray assistant. A glance at the apparatus and equipment in the background to which a seeming chaos of cables, pipes and houses are attached, are reminiscent of an intensive care unit. And yet Matthias Schmidt has never examined a patient – his test objects are fish. For his doctorate, he is researching into how warming and acidification of the oceans impact Arctic and Antarctic fish species. 

His special interest is focused on the fish brain. Australian scientists determined three years ago that young clown fish did not flee in acidic water but swam straight towards their enemy. In subsequent  tests, where the water was acidified, the juvenile fish were careless along their home reef, removed themselves further from their hideaways, saw and heard less well and within a few days over 70 per cent of the animals had been eaten. Matthias Schmidt now wishes to investigate whether fish in the polar regions react in a similar manner to ocean acidification with the assistance of NMR which is known as an MRT in hospitals.

The NMR system in its chamber
The NMR system in its chamber (Photo: Sina Löschke)

The NMR offers scientists many advantages. One of the greatest is that the animal can be examined “in vivo”, i.e. alive. Biologists such as Matthias Schmidt can therefore observe in real time how the acidic water and rising temperatures impact a fish through into the cells. They concentrate for example on the blood flow, i.e. the speed at which the heart pumps blood through the body. “Nobody really knows what acidification triggers in the brains of fish," explains Matthias Schmidt. He is looking for answers to questions that so far no one has asked. The research area the scientist dares to enter is therefore highly topical.

He is very well equipped for this challenge in the NMR laboratory of the Alfred Wegener Institute. The existing NMR equipment with a magnetic field strength of 4.7 Tesla exceeds the Earth’s magnetic field by approx. 100,000 times. The resolution of the image required by the scientists for their studies basically results from this field strength. By comparison, the MRT in hospitals usually reach strengths of between one and three Tesla for routine examinations on patients. However, the trend is towards ever larger magnets so that magnetic field strengths of 9.4 Tesla are no longer rare in experimental medical research.

Only a few research institutions have comparable investigation possibilities. But the real finesse of the system is the so-called swim tunnel. The “in vivo” NMR research group developed and built it together with the workshop of the Alfred Wegener Institute. This worldwide unique channel permits Matthias Schmidt to set different water speeds and observe the fish directly in the NMR. He can therefore conduct a kind of fitness test with the fish: the faster the water flows through the tunnel, the faster the fish has to swim. A separate feed and extraction system also enables him to regulate the carbon dioxide concentration in the water. Instead of increasing the resistance as on a treadmill, the biologist opens the carbon tap and makes the water more acidic. 

In this way, Matthias Schmidt can examine whether the carbon dioxide content in the water influences how fast the fish can swim. “The faster the fish swims, the faster the heart must also pump the blood to the brain. We suspect that fish in acidic water reach their physical limits more quickly," he explains.

warning against strong magnetic fields
Warning against strong magnetic fields (Photo: Sina Loeschke)

The biologist walks toward a chamber the size of a garage in the centre of the room and glances inside. This is where the NMR system is. The device looks like the MRT apparatus known in hospitals but the diameter of the examination tube is smaller. No human would fit inside. The magnetic coil of the NMR unit is located in a beige-coloured cylindrical housing. It floats here in liquid helium which in its turn is surrounded by liquid nitrogen and finally a vacuum. “The liquid gases cool the magnetic coil to approximately minus 270 Grad Celsius, i.e. just under absolute zero. Only then does the so-called supra conductivity occur in some metals, i.e. a current line virtually without resistance. Only in this way can a magnetic field of this kind be generated," explains Matthias Schmidt.



Matthias Schmidt regulates the water temperature and carbon dioxide concentration.
Matthias Schmidt regulates the water temperature and carbon dioxide concentration. (Photo: Sina Loeschke)
The water is cooled down to the right temperature.
The water is cooled down to the right temperature. (Photo: Sina Loeschke)
Examination tube of the AWI NMR
Examination tube of the NMR (Photo: Sina Löschke)
Closing the NMR camber
Closing the NMR camber (Photo: Sina Löschke)
Matthias Schmidt views the images from the NMR on the computer screen.
Matthias Schmidt views the images from the NMR on the computer screen. (Photo: Sina Loeschke)

His current test subject is an Antarctic Black Rockcod. The day before the biologist collected the fish from the Institute’s aquarium and brought it into the laboratory in a cold box filled with water. The biologist firstly had to slightly stun the animal because if the fish struggles it delays not only the experiment but primarily damages itself. After sedation, Matthias Schmidt had ten minutes to position his test subject such that it can survive experiments in the chamber over several days well and such that the scientist gets the best possible images. Precisely polar fish have such a small energy requirement that the animals can easily survive for several days without food.

When the fish comes around it finds itself in a dark container at zero degrees Celsius. Matthias Schmidt gives the fish twelve hours to become accustomed to its new environment. “We try to offer the fish conditions which are as similar to its usual environment as possible. In this case, this means that the fish should feel as comfortable as in our aquarium. After twelve hours, we can assume that it is no longer in a stressed state and only then do we start our experiments," says the biologist.

He closes the door to the chamber and sits down at the computer. From here he can observe precisely what is happening in the NMR and how the test subject is. Then he starts the first test. The NMR starts to knock and hammer. The chamber dampens the sound for those outside. But what does the fish feel like in the tube? “We have no indication that the fish is stressed. We can neither observe that it breathes quicker nor that it moves more. All records would lead us to believe that the noises do not cause any physiological stress with the fish," explains Matthias Schmidt pointing to his displays on the screen.

NMR lab at the Alfred Wegener Institute
Fish brain - a view inside (Photo: Sina Loeschke)
Aquaria at the Alfred Wegener Institute
Aquaria at the Alfred Wegener Institute (Photo: Kristina Bär)
Matthias Schmidt catches his proband out of the aquarium.
Matthias Schmidt catches his proband out of the fish tank. (Photo: Kristina Bär)

The first black and white specimen image of the fish brain appears on the computer screen. Matthias Schmidt is satisfied. “In the centre of the image we can see the brain. The dark areas signalise that the blood is flowing more slowly here and the lighter an area is, the faster the blood is flowing.” The biologist will be making just under 1500 measurements with the Black Rockcod and will analyse the results to determine how the fish reacted to the higher carbon dioxide concentrations in the water. 3D software will help him to analyse the recordings made. It simulates the brain of the fish and permits the biologist once again to take a closer look at any possible changes. The fish finally returns to the aquarium after the test where it is fed and where the next test subject is already waiting for Matthias Schmidt.
(Kristina Bär)