Marco Mank
Max Planck Society
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Featured researches published by Marco Mank.
Nature Methods | 2008
Marco Mank; Alexandre Ferrão Santos; Stephan Direnberger; Thomas D. Mrsic-Flogel; Sonja B. Hofer; Valentin Stein; Thomas Hendel; Dierk F. Reiff; Christiaan N. Levelt; Alexander Borst; Tobias Bonhoeffer; Mark Hübener; Oliver Griesbeck
Neurons in the nervous system can change their functional properties over time. At present, there are no techniques that allow reliable monitoring of changes within identified neurons over repeated experimental sessions. We increased the signal strength of troponin C–based calcium biosensors in the low-calcium regime by mutagenesis and domain rearrangement within the troponin C calcium binding moiety to generate the indicator TN-XXL. Using in vivo two-photon ratiometric imaging, we show that TN-XXL exhibits enhanced fluorescence changes in neurons of flies and mice. TN-XXL could be used to obtain tuning curves of orientation-selective neurons in mouse visual cortex measured repeatedly over days and weeks. Thus, the genetically encoded calcium indicator TN-XXL allows repeated imaging of response properties from individual, identified neurons in vivo, which will be crucial for gaining new insights into cellular mechanisms of plasticity, regeneration and disease.
Chemical Reviews | 2008
Marco Mank; Oliver Griesbeck
One of the greatest challenges in neuroscience has been to monitor electrical activity and biochemistry in populations of identified neurons in ViVo. Recent work on new microscopy techniques has moved the field considerably further in that direction. In particular the combination of modern imaging technology and genetic labeling methods heralds a bright future for neuronal circuit analysis. Biosensor development complements these efforts on the “indicator side” by providing probes for key events crucial for an understanding of neuronal function and plasticity. Moreover it aims at overcoming long-standing limitations in the ability to monitor neuronal activity and biochemistry in intact tissues and over longer periods of time. The term “genetically encoded” refers to the fact that the sensors are solely composed of amino acids with no addition of any synthetic compound or cofactor that would be difficult to apply into a living brain. Thus, sensors are encoded by a stretch of DNA that can be manipulated, mutated, and concatenated by any technique that the recombinant DNA toolbox offers. When DNA coding for the sensor is delivered into a cell, the sensor is formed within the cell in situ. Combined with the use of cell-type specific promoters, cellular targeting sequences, and transgenic technology, this offers a noninvasive means to implant an indicator deeply within a tissue of a living organism with cellular and subcellular specificity. Genetically encoded indicators use the Green Fluorescent Protein (GFP) or more generally speaking autofluorescent proteins (XFPs) and mutants thereof as fluorophores with all the advantages and restrictions that these fascinating proteins possess. Currently the XFPs are the only known family of proteins that form an internal fluorophore in aerobic environments via an autocatalytic reaction involving a number of critical amino acid residues within the peptide backbone. Therefore progress on the biosensor side is closely linked with further improvements in engineering XFPs for more brightness, photostability, full maturation at 37 °C, and shifts to longer wavelengths of emission. In its strictest sense the term “genetically encoded calcium indicators” (GECIs) also includes other types of sensors that operate with chemiluminescence, such as aequorin and derived sensors that use chemiluminescence energy transfer from aequorin to GFP. While offering some advantages compared to fluorescent reporters such as lack of background fluorescence and the ease of working with freely moving animals, the difficulty of obtaining high spatial resolution makes these sensors less suitable for many brain imaging applications. As the topic is relatively large, these probes will not be covered here in more detail. Instead the reader is invited to consult other reviews. Traditionally optical measurements of free cytosolic calcium fluctuations have been perfomed using a collection of synthetic organic molecules that change fluorescence or absorbance properties upon calcium binding. The field was revolutionized by the work of Roger Y. Tsien, who generated a series of fluorescent polycarboxylate compounds with strongly improved fluorescence properties. Both singlewavelength indicators and indicators suitable for ratiometric imaging were established. Tsien also introduced noninvasive methods of loading the compounds into living cells as acetoxymethyl esters. During the last two decades improvements in design and performance were incorporated. Currently, the best dyes are characterized by large fractional fluorescence changes (for example an approximately 14-fold relative intensity change for the indicator Oregon Green BAPTA-1; www.invitrogen.com), very good selectivity for calcium versus other cations, fast binding and dissociation kinetics, fairly linear response properties, pH-resistance, and photostability. Thus, synthetic fluorescent calcium probes provide a standard of performance parameters that genetically encoded calcium indicators will be compared with. Measurements of free cellular calcium have received special interest in the neurosciences, because the geometry and high degree of compartmentalization of neurons, together with an exquisite collection of calcium channels specific to neurons, has added another twist to the already ravishing complexity of cellular calcium handling and signaling. Moreover, rises in cytosolic calcium can be used as a relatively direct measure of neuronal activity. When a neuron fires action potentials, voltage-gated calcium channels in the plasma membrane open up and lead to a rise in cytosolic calcium within a few milliseconds. Due to the relatively large * To whom correspondence should be addressed. E-mail: griesbeck@ neuro.mpg.de. Chem. Rev. 2008, 108, 1550–1564 1550
Nature Methods | 2007
Nicola Heim; Olga Garaschuk; Michael W. Friedrich; Marco Mank; Ruxandra I Milos; Yury Kovalchuk; Arthur Konnerth; Oliver Griesbeck
Fluorescent Ca2+ indicator proteins (FCIPs) are attractive tools for studying Ca2+ dynamics in live cells. Here we describe transgenic mouse lines expressing a troponin C (TnC)-based biosensor. The biosensor is widely expressed in neurons and has improved Ca2+ sensitivity both in vitro and in vivo. This allows FCIP-based two-photon Ca2+ imaging of distinct neurons and their dendrites in vivo, and opens a new avenue for structure-function analysis of intact neuronal circuits.
The Journal of Neuroscience | 2008
Thomas Hendel; Marco Mank; Bettina Schnell; Oliver Griesbeck; Alexander Borst; Dierk F. Reiff
Recent advance in the design of genetically encoded calcium indicators (GECIs) has further increased their potential for direct measurements of activity in intact neural circuits. However, a quantitative analysis of their fluorescence changes (ΔF) in vivo and the relationship to the underlying neural activity and changes in intracellular calcium concentration (Δ[Ca2+]i) has not been given. We used two-photon microscopy, microinjection of synthetic Ca2+ dyes and in vivo calibration of Oregon-Green-BAPTA-1 (OGB-1) to estimate [Ca2+]i at rest and Δ[Ca2+]i at different action potential frequencies in presynaptic motoneuron boutons of transgenic Drosophila larvae. We calibrated ΔF of eight different GECIs in vivo to neural activity, Δ[Ca2+]i, and ΔF of purified GECI protein at similar Δ[Ca2+] in vitro. Yellow Cameleon 3.60 (YC3.60), YC2.60, D3cpv, and TN-XL exhibited twofold higher maximum ΔF compared with YC3.3 and TN-L15 in vivo. Maximum ΔF of GCaMP2 and GCaMP1.6 were almost identical. Small Δ[Ca2+]i were reported best by YC3.60, D3cpv, and YC2.60. The kinetics of Δ[Ca2+]i was massively distorted by all GECIs, with YC2.60 showing the slowest kinetics, whereas TN-XL exhibited the fastest decay. Single spikes were only reported by OGB-1; all GECIs were blind for Δ[Ca2+]i associated with single action potentials. YC3.60 and D3cpv tentatively reported spike doublets. In vivo, the KD (dissociation constant) of all GECIs was shifted toward lower values, the Hill coefficient was changed, and the maximum ΔF was reduced. The latter could be attributed to resting [Ca2+]i and the optical filters of the equipment. These results suggest increased sensitivity of new GECIs but still slow on rates for calcium binding.
Nature Neuroscience | 2010
Quoc Thang Nguyen; Lee F. Schroeder; Marco Mank; Arnaud Muller; Palmer Taylor; Oliver Griesbeck; David Kleinfeld
Tools from molecular biology, combined with in vivo optical imaging techniques, provide new mechanisms for noninvasively observing brain processes. Current approaches primarily probe cell-based variables, such as cytosolic calcium or membrane potential, but not cell-to-cell signaling. We devised cell-based neurotransmitter fluorescent engineered reporters (CNiFERs) to address this challenge and monitor in situ neurotransmitter receptor activation. CNiFERs are cultured cells that are engineered to express a chosen metabotropic receptor, use the Gq protein–coupled receptor cascade to transform receptor activity into a rise in cytosolic [Ca2+] and report [Ca2+] with a genetically encoded fluorescent Ca2+ sensor. The initial realization of CNiFERs detected acetylcholine release via activation of M1 muscarinic receptors. We used chronic implantation of M1-CNiFERs in frontal cortex of the adult rat to elucidate the muscarinic action of the atypical neuroleptics clozapine and olanzapine. We found that these drugs potently inhibited in situ muscarinic receptor activity.
Nature Neuroscience | 2010
Dierk F. Reiff; Johannes Plett; Marco Mank; Oliver Griesbeck; Alexander Borst
In the visual system of Drosophila, photoreceptors R1–R6 relay achromatic brightness information to five parallel pathways. Two of them, the lamina monopolar cells L1 and L2, represent the major input lines to the motion detection circuitry. We devised a new method for optical recording of visually evoked changes in intracellular Ca2+ in neurons using targeted expression of a genetically encoded Ca2+ indicator. Ca2+ in single terminals of L2 neurons in the medulla carried no information about the direction of motion. However, we found that brightness decrements (light-OFF) induced a strong increase in intracellular Ca2+ but brightness increments (light-ON) induced only small changes, suggesting that half-wave rectification of the input signal occurs. Thus, L2 predominantly transmits brightness decrements to downstream circuits that then compute the direction of image motion.
Nature Communications | 2012
Lai Hock Tay; Ivy E. Dick; Wanjun Yang; Marco Mank; Oliver Griesbeck; David T. Yue
Coupling of excitation to secretion, contraction, and transcription often relies upon Ca2+ computations within the nanodomain—a conceptual region extending tens of nanometers from the cytoplasmic mouth of Ca2+ channels. Theory predicts that nanodomain Ca2+ signals differ vastly from the slow submicromolar signals routinely observed in bulk cytoplasm. However, direct visualization of nanodomain Ca2+ far exceeds optical resolution of spatially distributed Ca2+ indicators. Here we couple an optical genetically encoded Ca2+ indicator (TN-XL) to the carboxyl tail of CaV2.2 Ca2+ channels, enabling nearfield imaging of the nanodomain. Under TIRF microscopy, we detect Ca2+ responses indicative of large-amplitude pulses. Single-channel electrophysiology reveals a corresponding Ca2+ influx of only 0.085 pA, and FRET measurements estimate TN-XL distance to the cytoplasmic mouth at ~55 Å. Altogether, these findings raise the possibility that Ca2+ exits the channel through the analog of molecular portals, mirroring the crystallographic images of side windows in voltage-gated K channels.
Journal of Biological Chemistry | 2010
Michael W. Friedrich; Gayane Aramuni; Marco Mank; Jonathan A. G. Mackinnon; Oliver Griesbeck
The Ca2+- and cAMP-responsive element-binding protein (CREB) and the related ATF-1 and CREM are stimulus-inducible transcription factors that link certain forms of cellular activity to changes in gene expression. They are attributed to complex integrative activation characteristics, but current biochemical technology does not allow dynamic imaging of CREB activation in single cells. Using fluorescence resonance energy transfer between mutants of green fluorescent protein we here develop a signal-optimized genetically encoded indicator that enables imaging activation of CREB due to phosphorylation of the critical serine 133. The indicator of CREB activation due to phosphorylation (ICAP) was used to investigate the role of the scaffold and anchoring protein AKAP79/150 in regulating signal pathways converging on CREB. We show that disruption of AKAP79/150-mediated protein kinase A anchoring or knock-down of AKAP150 dramatically reduces the ability of protein kinase A to activate CREB. In contrast, AKAP79/150 regulation of CREB via L-type channels may only have minor importance. ICAP allows dynamic and reversible imaging in living cells and may become useful in studying molecular components and cell-type specificity of activity-dependent gene expression.
Biophysical Journal | 2006
Marco Mank; Dierk F. Reiff; Nicola Heim; Michael W. Friedrich; Alexander Borst; Oliver Griesbeck
Georgia World Congress Center, Atlanta | 2006
Stephan Meyer zum Alten Borgloh; Ying Yang; Simone Astori; Peixin Zhu; Andrea Migala; Takeharu Nagai; Atsushi Miyawaki; Marco Mank; Oliver Griesbeck; Amy E. Palmer; Roger Y. Tsien; J. Nakai; Winfried Denk; Rolf Sprengel; Mazahir T. Hasan