Tracking neuronal activity in the living brain

Oct 22, 2010
Figure 1: The fluorescent indicator YC-Nano reveals waves of calcium flux (left) corresponding to signals generated by Dictyostelium cells as they undergo aggregation and action potentials in mouse cortical pyramidal cells (center) with a better signal to noise ratio (right). Credit: 2010 Katsuhiko Mikoshiba & Takeharu Nagai

Refinements to a fluorescent calcium ion indicator give scientists a powerful tool for tracking neuronal activity in the living brain

As travel along chains of neurons, each cell undergoes a dramatic shift in its internal calcium ion (Ca2+) concentration because specialized channels allow to flood into the . This shift provides a valuable indicator for tracking neural activity in real time, so scientists have developed several fluorescent protein-based Ca2+ indicators that are genetically encoded and can therefore be expressed directly in of interest.

Generally these indicators do not perform as well in live animals as in vitro. Takeharu Nagai of Hokkaido University and Katsuhiko Mikoshiba of the RIKEN Brain Science Institute in Wako suspected that indicators with higher affinity for Ca2+ might work better. However, their approach was risky. “It was generally believed that extremely high-affinity Ca2+ indicators would result in low cell viability due to disturbed Ca2+ homeostasis, and show no signal changes due to saturation by resting Ca2+,” say Nagai and Mikoshiba. “From this point of view, our attempt was totally against common sense.”

Nevertheless, the indicators, dubbed YC-Nano, developed by Nagai and his colleagues proved to be a remarkable success. The indicators were derived from yellow cameleon (YC), a genetically encoded indicator consisting of two fluorescent proteins, a ‘donor’ and an ‘acceptor’, connected by a Ca2+-binding domain. In the presence of Ca2+, the structure of YC rearranges such that the two come close together in a manner that allows energy from the excited donor to induce a readily detectable signal from the acceptor; in the absence of Ca2+, only a minimal signal is produced.

The researchers introduced various modifications that lengthened the Ca2+-binding segment between the two fluorescent domains, introducing additional flexibility that considerably improved indicator sensitivity. The best-performing versions exhibited five-fold greater Ca2+ affinity than YC and a high dynamic range. “We were quite surprised that we managed to systematically produce a series of indicator variants with different affinity by a very simple protein engineering trick,” says Nagai.

YC-Nano accurately tracked the complex patterns of Ca2+ activation seen in the aggregating process of social amoeba Dictyostelium, revealing propagating waves throughout the aggregates in a rotating spiral. These indicators also performed well in monitoring in the brains of mice, and Mikoshiba foresees numerous experimental applications in the near future. “Since YC-Nano can be stably expressed in specific types of neurons for a long range of time,” he says, “we expect to perform chronic in vivo imaging and analyze the modifications of neuronal network activities underlying learning, development or diseases of the brain.”

Explore further: REM sleep critical for young brain development; medication interferes

More information: Horikawa, K., et al. Spontaneous network activity visualized by ultrasensitive Ca2+ indicators, yellow Cameleon-Nano. Nature Methods 7, 729–732 (2010).

Related Stories

A warm sensor maintains skin barrier

May 14, 2010

Japanese research group led by Prof. Makoto Tominaga and Dr. Takaaki Sokabe (National Institute for Physiological Sciences: NIPS) found that TRPV4 ion channel in skin keratinocytes is important for formation and maintenance ...

Gene limits learning and memory in mice

Sep 17, 2010

Deleting a certain gene in mice can make them smarter by unlocking a mysterious region of the brain considered to be relatively inflexible, scientists at Emory University School of Medicine have found.

New perspectives on local calcium signaling

Jul 26, 2010

The latest Perspectives in General Physiology series introduces the newest technologies in the field of calcium signaling, which plays a central role in many cellular processes. The Perspectives appear in ...

What causes seizure in focal epilepsy?

Apr 13, 2010

In focal epilepsy, seizures are generated by a localized, synchronous neuronal electrical discharge that may spread to large portions of the brain. In spite of intense research in the field of epilepsy, a key question remains ...

Recommended for you

Making waves with groundbreaking brain research

19 hours ago

New research by Jason Gallivan and Randy Flanagan suggests that when deciding which of several possible actions to perform, the human brain plans multiple actions simultaneously prior to selecting one of ...

Long-term memories are maintained by prion-like proteins

Jul 02, 2015

Research from Eric Kandel's lab at Columbia University Medical Center (CUMC) has uncovered further evidence of a system in the brain that persistently maintains memories for long periods of time. And paradoxically, ...

Water to understand the brain

Jul 02, 2015

To observe the brain in action, scientists and physicians use imaging techniques, among which functional magnetic resonance imaging (fMRI) is the best known. These techniques are not based on direct observations ...

User comments : 0

Please sign in to add a comment. Registration is free, and takes less than a minute. Read more

Click here to reset your password.
Sign in to get notified via email when new comments are made.