Neurons and Muscle Cells Need Stabilizing Force for Effective Communication

Neurons and Muscle Cells Need Stabilizing Force for Effective Communication Thursday, 09 October 2008 You cannot raise a finger without your brain directing muscle cells, and scientists have figured out another reason that usually works so well. A neuron sends a message, or neurotransmitter, to the muscle cell to tell it what to do. To get the message, the receiving cell must have a receptor. Oddly, the unstable protein rapsyn is responsible for anchoring the receptor so it is properly positioned to catch the message. Medical College of Georgia scientists have found what keeps rapsyn in proper conformation. It is a heat shock protein, one of a large family of molecular chaperones that make sure proteins get where they are needed and do what they should, says Dr. Lin Mei, chief of developmental neurobiology at MCG and Georgia Research Alliance Eminent Scholar in Neuroscience. Hsp90β helps stabilize rapsyn so receptors can get and stay where needed, according to research published in the Oct. 9 issue of Neuron. Dr. Mei suspects that other hsp siblings have a similar caretaker role in neuron-to-neuron communication in the brain. Scientists knew rapsyn's role in getting neuromuscular receptors to aggregate and stay where needed, but they didn't know what stabilized it. "It makes you wonder how to control this naughty boy which is very important," says Dr. Mei, the study's corresponding author. They found hsp90β wherever rapsyn clustered in muscle cells. When they disrupted its activity or expression, they realized hsp90β's stabilizing role in forming and maintaining receptor clusters, says Dr. Shiwen Luo, postdoctoral fellow in Dr. Mei's lab and the study's first author. Rapsyn and the receptor apparently interact, and then hsp90β comes along to help stabilize the relationship. Rapsyn mutations have been implicated in muscular dystrophies including congenital myasthenia gravis. MCG researchers are looking now to see if a mutated rapsyn still interacts with hsp90β. They used a type of acetylcholine nicotinic receptor at the neuromuscular junction as a model for their studies of brain development and communication. The junction is 1,000 times larger than connections, or synapses, between two neurons but structurally similar. Fundamentals include presynaptic terminals that release neurotransmitters picked up by receptors on the postsynaptic side. Terminals and receptors must be lined up well, whether it's a muscle cell or neuron getting the message. "In central nervous system synapses and at the neuromuscular junction, receptors have to be concentrated at the right spot to receive the neurotransmitter released," says Dr. Mei. If receptors are in the wrong place, the message can be weak or even lost. At the neuromuscular juncture, communication is usually straightforward, with primarily one neurotransmitter and one principal receptor. "Whenever you tell a muscle to move, it moves. If you want your muscles to think, you wouldn't be able to pick up a pin," says Dr. Mei. In the brain, where neurons have thousands of synapses, it is more of a negotiation. "Signals have to be integrated in the neuron for it to decide what to do." ......... ZenMaster

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Artificial Cells Could Power Medical Implants

Models of eel cells suggest electrifying possibilities Thursday, 09 October 2008 Researchers at Yale University have created a blueprint for artificial cells that are more powerful and efficient than the natural cells they mimic and could one day be used to power tiny medical implants. The scientists began with the question of whether an artificial version of the electrocyte – the energy-generating cells in electric eels – could be designed as a potential power source. "The electric eel is very efficient at generating electricity," said Jian Xu, a postdoctoral associate in Yale's Department of Chemical Engineering. "It can generate more electricity than a lot of electrical devices." Xu came up with the first blueprint that shows how the electrocyte's different ion channels work together to produce the fish's electricity while he was a graduate student under former Yale assistant professor of mechanical engineering David LaVan, now at the National Institute of Standards and Technology. But the scientists didn't stop there. "We're still trying to understand how the mechanisms in these cells work," said LaVan. "But we asked ourselves: 'Do we know enough to sit down and start thinking about how to build these things’ — and to optimize that design? Nobody had really done that before." Using the new blueprint as a guide, LaVan and Xu set about designing an artificial cell that could replicate the electrocyte's energy production. "We wanted to see if nature had already optimized the power output and energy conversion efficiency of this cell," said Xu. "And we found that an artificial cell could actually outperform a natural cell, which was a very surprising result." The artificial cell LaVan and Xu modelled is capable of producing 28 percent more electricity than the eel's own electrocyte, with 31 percent more efficiency in converting the cell's chemical energy – derived from the eel's food – into electricity. Electric eels channel the output of thousands of specialized cells called electrocytes to generate electric potentials of up to 600 volts, according to biologists. The mechanism is similar to nerve cells. The arrival of a chemical signal triggers the opening of highly selective channels in a cell membrane causing sodium ions to flow in and potassium ions to flow out. The ion swap increases the voltage across the membrane, which causes even more channels to open. Past a certain point the process becomes self-perpetuating, resulting in an electric pulse travelling through the cell. The channels then close and alternate paths open to “pump” the ions back to their initial concentrations during a “resting” state.

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Electric eel anatomy. The first detail shows stacks of electrocytes, cells linked in series (to build up voltage) and parallel (to build up current). Second detail shows an individual cell with ion channels and pumps penetrating the membrane, The Yale/NIST model represents the behaviour of several such cells. Final detail shows an individual ion channel, one of the building blocks of the model. Credit: Daniel Zukowski, Yale University

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In all, according LaVan, there are at least seven different types of channels, each with several possible variables to tweak, such as their density in the membrane. Nerve cells, which move information rather than energy, can fire rapidly but with relatively little power. Electrocytes have a slower cycle, but deliver more power for longer periods. LaVan and Xu developed a complex numerical model to represent the conversion of ion concentrations to electrical impulses and tested it against previously published data on electrocytes and nerve cells to verify its accuracy. Then they considered how to optimize the system to maximize power output by changing the overall mix of channel types. Their calculations show that substantial improvements are possible. One design for an artificial cell generates more than 40 percent more energy in a single pulse than a natural electrocyte. Another would produce peak power outputs over 28 percent higher. In principle, say the authors, stacked layers of artificial cells in a cube slightly over 4 mm on a side are capable of producing continuous power output of about 300 microwatts to drive small implant devices. The individual components of such artificial cells — including a pair of artificial membranes separated by an insulated partition and ion channels that could be created by engineering proteins — already have been demonstrated by other researchers. Like the natural counterpart, the cell’s energy source would be adenosine triphosphate (ATP), synthesized from the body’s sugars and fats using tailored bacteria or mitochondria. While eels use thousands of electrocytes to produce charges of up to 600 volts, LaVan and Xu’s calculations show it would be possible to create a smaller "bio-battery" using several dozen artificial cells. The tiny bio-batteries would only need to be about 4 mm thick to produce the small voltages needed to power tiny electrical devices such as retinal implants or other prostheses. Although the engineers came up with a design, it will still be some time before the artificial cells are actually built. For one thing, they still need a power source before they could start producing electricity. LaVan speculates the cells could be powered in a way similar to their natural counterparts. It is possible, he said, that bacteria could be employed to recycle ATP – responsible for transferring energy within the cell – using glucose, a common source of chemical energy derived from food. With an energy source in place, the artificial cells could one day power medical implants and would provide a big advantage over battery-operated devices. "If it breaks, there are no toxins released into your system," said Xu. "It would be just like any other cell in your body." Reference: Designing artificial cells to harness the biological ion concentration gradient J. Xu and D.A. LaVan. Nature Nanotechnology advanced online publication 10.1038/nnano.2008.274 ......... ZenMaster

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Living Cells as Nanotechnology Factories

Bacteria used as factories to make complex DNA based nanostructures Thursday, 09 October 2008 In the tiny realm of nanotechnology, scientists have used a wide variety of materials to build atomic scale structures. But just as in the construction business, nanotechnology researchers can often be limited by the amount of raw materials. Now, Biodesign Institute at Arizona State University researcher Hao Yan has avoided these pitfalls by using cells as factories to make DNA based nanostructures inside a living cell. Yan specializes in a fast-growing field within nanotechnology — commonly known as structural DNA nanotechnology — that uses the basic chemical units of DNA, abbreviated as C, T, A, or G, to self-fold into a number of different building blocks that can further self-assemble into patterned structures. "This is a good example of artificial nanostructures that can be replicated using the machineries in live cells" said Yan. "Cells are really good at making copies of double stranded DNA and we have used the cell like a copier machine to produce many, many copies of complex DNA nanostructures." DNA nanotechnologists have made some very exciting achievements during the past five to 10 years. Nevertheless, DNA nanotechnology has been limited by the need to chemically synthesize all of the material from scratch. To date, it has strictly been a test tube science, where researchers have developed many toolboxes for making different DNA nanostructures to attach and organize other molecules including nanoparticles and other biomolecules. "If you need to make a single gram of a DNA nanostructure, you need to order one gram of the starting DNA materials. Scientists have previously used chemical methods to copy branched DNA structures, and there has also been significant work in using long-stranded DNA sequences replicated from cells or phage viruses to scaffold short helper DNA sequences to form 2-D or 3-D objects," said Yan, who is a professor in the Department of Chemistry and Biochemistry at ASU. "We have always dreamed of scaling up DNA nanotechnology. One way to scale it up is to use the cellular system because simple DNA can be replicated inside the cell. We wanted to know if the cell's copy machine could tolerate single stranded DNA nanostructures that contain complicated secondary structures." Yan and his fellow researchers, Chenxiang Lin, Sherri Rinker and Yan Liu at ASU and their collaborators Ned Seeman and Xing Wang at New York University, wanted to test the nanoscale manufacturing capabilities of cells. They went back to reproducing the very first branched nanostructure made up of DNA — a cross-shaped, four-arm DNA junction. Another DNA junction structure containing a different crossover topology was also tested. To copy these branched DNA nanostructures inside a living cell, the ASU and NYU research team first shipped the cargo inside a bacterial cell. They cut and pasted the DNA necessary to make these structures into a phagemid, a virus-like particle that infects a bacterial cell. Once inside the cell, the phagemid used the cell just like a photocopier machine to reproduce millions of copies of the DNA. By theoretically starting with just a single phagemid infection, and a single millilitre of cultured cells, Yan found that the cells could churn out trillions of the DNA junction nanostructures. The DNA nanostructures produced in the cells were also found to fold correctly, just like the previously built test tube structures. According to Yan, the results proved the key existence of the DNA nanostructures during the cell's routine DNA replication and division cycles. "When a DNA nanostructure gets replicated, it does exist and can survive the complicated cellular machinery. And it looks like the cell can tolerate this kind of structure and still do its job. It's amazing," Yan said. Yan acknowledges that this is just the first step, but foresees there are many interesting DNA variations to consider next. "The fact that the natural cellular machinery can tolerate artificial DNA objects is quite intriguing, and we don't know what the limit is yet." Yan's group may be able to change and evolve DNA nanostructures and devices using the cellular system and the technology may also open up some possibilities for synthetic biology applications. "I'm very excited about the future of DNA nanotechnology, but there is a lot of work to be done. An interesting research topic to pursue is the interface of DNA nanostructures with live cells; it is full of opportunities," said Yan. The results were published in the early online edition of the Proceedings of the National Academy of Sciences. ......... ZenMaster

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