.<\/p>\n
As electrical signals travel along chains of neurons, each cell undergoes
\n a dramatic shift in its internal calcium ion (Ca2+) concentration because
\nspecialized channels allow ions to flood into the cytoplasm. This shift
\nprovides a valuable indicator for tracking neural activity in real time, so
\n scientists have developed several fluorescent protein-based Ca2+
\nindicators that are genetically encoded and can therefore be expressed
\n directly in cells of interest.
\n.
\nGenerally these indicators do not perform as well in live animals as in vitro.
\nTakeharu Nagai of Hokkaido University and Katsuhiko Mikoshiba of the
\nRIKEN Institute, suspected that indicators with higher affinity for Ca2+ might
\nwork better. However, their approach was risky. \u201cIt was generally believed
\nthat extremely high-affinity Ca2+ indicators would result in low cell viability
\ndue to disturbed Ca2+ homeostasis, and show no signal changes due to
\n saturation by resting Ca2+,\u201d say Nagai and Mikoshiba. \u201cFrom this point of
\nview, our attempt was totally against common sense.\u201d
\n.
\nNevertheless, the indicators, dubbed YC-Nano, developed by Nagai and his
\ncolleagues proved to be a remarkable success1. The indicators were derived
\nfrom yellow cameleon (YC), a genetically encoded indicator consisting of two
\nfluorescent proteins, a \u2018donor\u2019 and an \u2018acceptor\u2019, connected by a Ca2+-binding
\ndomain. In the presence of Ca2+, the structure of YC rearranges such that the
\ntwo come close together in a manner that allows energy from the excited donor
\n to induce a readily detectable signal from the acceptor; in the absence
\nof Ca2+, only a minimal signal is produced.
\n.
\nThe researchers introduced various modifications that lengthened the Ca2+
\n-binding segment between the two fluorescent domains, introducing additional
\nflexibility that considerably improved indicator sensitivity. The best-performing
\nversions exhibited five-fold greater Ca2+ affinity than YC and a high dynamic
\nrange. \u201cWe were quite surprised that we managed to systematically produce a
\nseries of indicator variants with different affinity by a very simple protein
\nengineering trick,\u201d says Nagai.
\n.
\nYC-Nano accurately tracked the complex patterns of Ca2+ activation seen in the
\naggregating process of social amoeba Dictyostelium, revealing propagating
\n waves throughout the aggregates in a rotating spiral. These indicators
\n also performed well in monitoring neuronal activity in the brains of mice, and
\n Mikoshiba foresees numerous experimental applications in the near future.
\n\u201cSince YC-Nano can be stably expressed in specific types of neurons for a
\nlong range of time,\u201d he says, \u201cwe expect to perform chronic in vivo
\nimaging and analyze the modifications of neuronal network activities
\nunderlying learning, development or diseases of the brain.\u201d
\n.<\/p>\n","protected":false},"excerpt":{"rendered":"
. As electrical signals travel along chains of neurons, each cell undergoes a dramatic shift in its internal calcium ion (Ca2+) concentration because specialized channels allow ions to flood into the cytoplasm. This shift provides a valuable indicator for tracking neural activity in real time, so scientists have developed several fluorescent protein-based Ca2+ indicators that … <\/p>\n