Current Topics in Medicinal Chemistry - Volume 16, Issue 24, 2016

Firefly shows bioluminescence by “luciferin−luciferase” (L−L) reaction using luciferin, luciferase, ATP and O2. The chemical photon generation by an enzymatic reaction is widely utilized for analytical methods including biological imaging in the life science fields. To expand photondetecting analyses with firefly bioluminescence, it is important for users to understand the chemical basis of the L−L reaction. In particular, the emission color variation of the L−L reaction is one of the distinguishing characteristics for multicolor luciferase assay and in vivo imaging. From the viewpoint of fundamental chemistry, this review explains the recent progress in the studies on the molecular mechanism of emission color variation after showing the outline of the reaction mechanism of the whole L−L reaction. On the basis of the mechanism, the progresses in organic synthesis of luciferin analogs modulating their emission colors are also presented to support further developments of red/near infrared in vivo biological imaging utility of firefly bioluminescence.

Firefly bioluminescence is widely used in life science research as a useful analysis tool. For example, the adenosine-5′-triphosphate (ATP)-dependent enzymatic firefly bioluminescence reaction has long been utilized as a microbial monitoring tool. Rapid and sensitive firefly luciferin-luciferase combinations are used not only to measure cell viability but also for reporter-gene assays. Recently, bioluminescence was utilized as a noninvasive, real-time imaging tool for living subjects to monitor cells and biological events. However, the number of commercialized luciferase genes is limited and tissue-permeable near-infrared (NIR) region emitting light is required for in vivo imaging. In this review, recent studies describing synthetic luciferin analogues predicted to have red-shifted bioluminescence are summarized. Luciferase substrates emitting red, green, and blue light that were designed and developed in our laboratory are presented. The longest emission wavelength of the synthesized luciferin analogues was recorded at 675 nm, which is within the NIR region. This compound is now commercially available as “Aka Lumine®”.

Skeletal muscle contractions and their attendant increase in metabolism accelerate production and accumulation of multiple metabolites in the cytoplasm, interstitium and vascular compartments. Continued myocyte contractile function and subsequent recovery is often highly dependent upon constraining metabolite accumulation. In particular, the control of pH ([H+]) during exercise is absolutely crucial for muscle force maintenance and metabolic regulation. This short review focuses attention on the dynamic control mechanisms that help regulate H+ and thus intramyocyte pH (pHi) during muscle contractions: Specifically 1. Myocyte H+ production and intrinsic buffering systems, 2. Mechanisms of myocyte H+ excretion, and 3. Inter-relationships among the intramyocyte - interstitial – blood [H+].

The innovation and development of live-cell fluorescence imaging methods have revealed the dynamic aspects of intracellular Ca2+ in a wide variety of cells. The fertilized egg, the very first cell to be a new individual, has long been under extensive investigations utilizing Ca2+ imaging since its early days, and spatiotemporal Ca2+ dynamics and underlying mechanisms of Ca2+ mobilization, as well as physiological roles of Ca2+ at fertilization, have become more or less evident in various animal species. In this article, we illustrate characteristic patterns of Ca2+ dynamics in mammalian gametes and molecular basis for Ca2+ release from intracellular stores leading to the elevation in cytoplasmic Ca2+ concentration, and describe the identity and properties of sperm-borne egg-activating factor in relation to the induction of Ca2+ waves and Ca2+ oscillations, referring to its potential use in artificial egg activation as infertility treatment. In addition, a possible Ca2+ influx-driven mechanism for slow and long-lasting Ca2+ oscillations characteristic of mammalian eggs is proposed, based on the recent experimental findings and mathematical modeling. Cumulative knowledge about the roles of Ca2+ in the egg activation leading to early embryogenesis is summarized, to emphasize the diversity of functions that Ca2+ can perform in a single type of cell.

An adequate supply of blood flow to the brain is critically important to maintain long-term brain function. However, many issues surrounding the regulatory mechanism of the blood flow supply to the brain remain unclear, such as i) the appropriate range of capillary flow velocity to keep neurons healthy, ii) the size of the vascular module to support a functioning neural unit, iii) the sensing mechanism for capillary flow control, and iv) the role of flow regulation in promoting neural plasticity. A fluorescence technique allows for visualization of the dynamic changes between cerebral microcirculation and neural activity concurrently and thus is capable of addressing these questions. Here, we briefly review the methodological aspects of measuring blood flow using fluorescence imaging in rodent brains and introduce a novel approach for mapping the flow velocity in multiple vessels with laser scanning fluorescence microscopy. The flow velocity was imaged by calculating the traveling distance and time of the instantaneously injected fluorescent tags through the vascular networks. Using the present method, we observed that the average flow velocity in the pial artery and vein was 3.0 ± 1.4 mm/s and 1.6 ± 0.5 mm/s, respectively (N = 6 mice). A limitation of the method presented is that the quantification is only applicable to the vascular networks laid in two-dimensional planes, such as pial vascular networks. Further technical improvement is needed to quantify three-dimensional flow through parenchymal microcirculation. Furthermore, it is also needed to fill a gap between the microscopically measured flow parameters and the macroscopic feature of the brain blood flow for clinical interpretation.

Magnetic resonance imaging (MRI) of the human brain plays an important role in the field of medical imaging as well as basic neuroscience. It measures proton spin relaxation, the time constant of which depends on tissue type, and allows us to visualize anatomical structures in the brain. It can also measure functional signals that depend on the local ratio of oxyhemoglobin to deoxyhemoglobin in the blood, which is believed to reflect the degree of neural activity in the corresponding area. MRI thus provides anatomical and functional information about the human brain with high spatial resolution. Conventionally, MRI signals are measured and analyzed for each individual voxel. However, these signals are essentially multivariate because they are measured from multiple voxels simultaneously, and the pattern of activity might carry more useful information than each individual voxel does. This paper reviews recent trends in multivariate analysis of MRI signals in the human brain, and discusses applications of this technique in the fields of medical imaging and neuroscience.

A radioisotope and its imaging have been powerful tools to explore the mechanism of chemical reaction and the dynamic behavior of trace element in biomolecular science and nuclear medicine. This article reviews some labelled compounds for radiopharmaceuticals in biochemical science and nuclear medicine using short-lived nuclides, the production and applications of “Multitracer” containing carrier-free radionuclides of more than 50 elements and a new radioactive tracer technique coupled with simultaneously imaging of multiple molecules labelled by γ-emitters. In addition, X-ray phase imaging and microscopy technique using synchrotron radiation are introduced.

Molecular imaging can be used to evaluate the spatial–time change of the molecular biological phenomenon of the cell–molecule level in living bodies. Molecular imaging technology is expected to be applied in the fields of drug development, clinical diagnosis, and life science research. Specifically, positron emission tomography (PET) is a powerful non-invasive imaging technology for investigating physiological parameters in living animals using compounds labeled with PET radioisotopes as molecular probes. This review summarizes and compares various 18F-conjugation techniques that employ the chemical and enzymatic reactions of different types of tumor-targeting biological molecules such as peptides, proteins, antibodies, and nucleic acids.