α7 nicotinic acetylcholine receptor upregulation by anti-apoptotic Bcl-2 proteins.

Nicotinic acetylcholine receptors (nAChRs) mediate and modulate synaptic transmission throughout the brain, and contribute to learning, memory, and behavior. Dysregulation of α7-type nAChRs in neuropsychiatric as well as immunological and oncological diseases makes them attractive targets for pharmaceutical development. Recently, we identified NACHO as an essential chaperone for α7 nAChRs. Leveraging the robust recombinant expression of α7 nAChRs with NACHO, we utilized genome-wide cDNA library screening and discovered that several anti-apoptotic Bcl-2 family proteins further upregulate receptor assembly and cell surface expression. These effects are mediated by an intracellular motif on α7 that resembles the BH3 binding domain of pro-apoptotic Bcl-2 proteins, and can be blocked by BH3 mimetic Bcl-2 inhibitors. Overexpression of Bcl-2 member Mcl-1 in neurons enhanced surface expression of endogenous α7 nAChRs, while a combination of chemotherapeutic Bcl2-inhibitors suppressed neuronal α7 receptor assembly. These results demonstrate that Bcl-2 proteins link α7 nAChR assembly to cell survival pathways.

NACHO Mediates Nicotinic Acetylcholine Receptor Function throughout the Brain.

Neuronal nicotinic acetylcholine receptors (nAChRs) participate in diverse aspects of brain function and mediate behavioral and addictive properties of nicotine. Neuronal nAChRs derive from combinations of α and ß subunits, whose assembly is tightly regulated. NACHO was recently identified as a chaperone for α7-type nAChRs. Here, we find NACHO mediates assembly of all major classes of presynaptic and postsynaptic nAChR tested. NACHO acts at early intracellular stages of nAChR subunit assembly and then synergizes with RIC-3 for receptor surface expression. NACHO knockout mice show profound deficits in binding sites for α-bungarotoxin, epibatidine, and conotoxin MII, illustrating essential roles for NACHO in proper assembly of α7-, α4ß2-, and α6-containing nAChRs, respectively. By contrast, GABAA receptors are unaffected consistent with NACHO specifically modulating nAChRs. NACHO knockout mice show abnormalities in locomotor and cognitive behaviors compatible with nAChR deficiency and underscore the importance of this chaperone for physiology and disease associated with nAChRs.

Optodynamic simulation of ß-adrenergic receptor signalling.

Optogenetics has provided a revolutionary approach to dissecting biological phenomena. However, the generation and use of optically active GPCRs in these contexts is limited and it is unclear how well an opsin-chimera GPCR might mimic endogenous receptor activity. Here we show that a chimeric rhodopsin/ß2 adrenergic receptor (opto-ß2AR) is similar in dynamics to endogenous ß2AR in terms of: cAMP generation, MAP kinase activation and receptor internalization. In addition, we develop and characterize a novel toolset of optically active, functionally selective GPCRs that can bias intracellular signalling cascades towards either G-protein or arrestin-mediated cAMP and MAP kinase pathways. Finally, we show how photoactivation of opto-ß2AR in vivo modulates neuronal activity and induces anxiety-like behavioural states in both fiber-tethered and wireless, freely moving animals when expressed in brain regions known to contain ß2ARs. These new GPCR approaches enhance the utility of optogenetics and allow for discrete spatiotemporal control of GPCR signalling in vitro and in vivo.

Ultraminiaturized photovoltaic and radio frequency powered optoelectronic systems for wireless optogenetics.

OBJECTIVE: Wireless control and power harvesting systems that operate injectable, cellular-scale optoelectronic components provide important demonstrated capabilities in neuromodulatory techniques such as optogenetics. Here, we report a radio frequency (RF) control/harvesting device that offers dramatically reduced size, decreased weight and improved efficiency compared to previously reported technologies. Combined use of this platform with ultrathin, multijunction, high efficiency solar cells allows for hundred-fold reduction of transmitted RF power, which greatly enhances the wireless coverage. APPROACH: Fabrication involves separate construction of the harvester and the injectable µ-ILEDs. To test whether the presence of the implantable device alters behavior, we implanted one group of wild type mice and compared sociability behavior to unaltered controls. Social interaction experiments followed protocols defined by Silverman et al. with minor modifications. MAIN RESULTS: The results presented here demonstrate that miniaturized RF harvesters, and RF control strategies with photovoltaic harvesters can, when combined with injectable µ-ILEDs, offer versatile capabilities in optogenetics. Experimental and modeling studies establish a range of effective operating conditions for these two approaches. Optogenetics studies with social groups of mice demonstrate the utility of these systems. SIGNIFICANCE: The addition of miniaturized, high performance photovoltaic cells significantly expands the operating range and reduces the required RF power. The platform can offer capabilities to modulate signaling path in the brain region of freely-behaving animals. These suggest its potential for widespread use in neuroscience.

11C-LY2428703, a positron emission tomographic radioligand for the metabotropic glutamate receptor 1, is unsuitable for imaging in monkey and human brains.

BACKGROUND: A recent study from our laboratory demonstrated that 11C-LY2428703, a new positron emission tomographic radioligand for metabotropic glutamate receptor 1 (mGluR1), has promising in vitro properties and excellent in vivo performance for imaging rat brain. The present study evaluated 11C-LY2428703 for imaging mGluR1 in monkey and human brains. METHODS: Rhesus monkeys were imaged at baseline and after administration of an mGluR1 blocking agent to calculate nonspecific binding, as well as after the administration of permeability glycoprotein (P-gp) and breast cancer resistance protein (BCRP) blockers to assess whether 11C-LY2428703 is a substrate for efflux transporters at the blood-brain barrier. Human imaging was performed at baseline in three healthy volunteers, and arterial input function was measured. RESULTS: Overall brain uptake was low in monkeys, though slightly higher in the cerebellum, where mGluR1s are concentrated. However, the uptake was not clearly displaceable in the scans after mGluR1 blockade. Brain penetration of the ligand did not increase after P-gp and BCRP blockade. Brain uptake was similarly low in all human subjects (mean VT with a two-tissue compartment model, 0.093 ± 0.012 mL/cm3) and for all regions, including the cerebellum. CONCLUSIONS: Despite promising in vitro and in vivo results in rodents, 11C-LY2428703 was unsuitable for imaging mGluR1s in monkey or human brain because of low brain uptake, which was likely caused by high binding to plasma proteins.