Channeling Neuroscience

A SHORT STINT IN MEDICINE

Although Jonas yielded to her family's wishes and enrolled in medical school at New York University, she also succeeded in pursuing a career in neuroscience. After completing her medical degree, Jonas returned to New Haven to do a residency in neurology. During her final year as a resident, she opted to do an elective in the laboratory of Len Kaczmarek at the Yale University School of Medicine.

While there, Jonas invented a technique for recording the activity of ion channels that adorn intracellular organelles inside living cells. This technological tour-de-force has allowed her to examine the role that mitochondria play in regulating neurotransmitter release and controlling whether individual synapses will survive or die. "She's studying a cell biological problem at the synapse," says Jennifer Morgan of the University of Texas in Austin. Jonas' work "puts her at the cutting edge because she's bridging these two fields of cell biology and neurobiology."

Because Jonas didn't follow the traditional route - that would have meant getting a PhD before doing a postdoc with Kaczmarek - her learning curve was steep. "I didn't even know how to make a solution," says Jonas, now an assistant professor of internal medicine at Yale. Never having worked at the bench, Jonas says she made plenty of rookie mistakes. Once during a lab meeting, for example, one of Kaczmarek's graduate students announced that someone had left the pH meter probe dangling in the air rather than immersed in buffer. "I knew I was the one," laughs Jonas. "That's how I was in the lab."

"Liz will say that she didn't know what she was doing for the first few years," says Kaczmarek. "Sure, it took her awhile to get to be the machine she is now, where she's just doing experiments every waking moment. But she was enthusiastic and she loved doing experiments."

Jonas still wasn't enamored with medicine. "She really hated it," says Kaczmarek. "Liz ran a headache clinic one day a week while she was in my lab. She would be in a really bad mood the day before and the day of the clinic. A really bad mood. When it was over, her mood would change completely. I think she's a good doctor, she's really caring with her patients, but it distracted her from what she really wants to do."

IN SEARCH OF CHANNELS

What Jonas really wanted to do was look for calcium channels that she thought should be located in the membranes of the neurotransmitter-containing vesicles that pack within axon terminals awaiting the signal to dump their chemical load into the synapse. Calcium is needed in the cytosol to allow the vesicles to fuse with the plasma membrane, and that calcium has to come from somewhere. Much comes in through the plasma membrane, but some comes from intracellular calcium stores, including, Jonas reasoned, the vesicles themselves. Then one afternoon she was listening to Barbara Ehrlich (also at Yale) lecture about recording the activity of the IP3 receptor, which acts as a calcium channel in the endoplasmic reticulum (ER) membrane. Ehrlich had reconstituted the IP3 channel in an artificial lipid bilayer.

"I sat there thinking it would be so nice to see that channel open inside a living cell," says Jonas, "and all of a sudden the way to do that just popped into my head." Jonas realized that to record channel activity from a membrane inside the cell, she needed to use two electrodes, one nestled inside the other. She would use the outer electrode to poke through the plasma membrane. Once both electrodes were inside the cell, she would then withdraw the outer one, which could be contaminated with bits of the plasma membrane. The clean inner electrode could then be used to patch on to whatever inner membrane she might encounter. "I went to Len thinking that this was a great idea, and Len said, ?you can't work on that. That is ridiculous.'"

Although Kaczmarek contends that he probably said something more tactful, like "it might not be such a good idea to spend too much time on this," he admits that he didn't hold out much hope for the project. "It seemed a little bit complicated, having one electrode inside another," says Kaczmarek. He wondered whether the cell would survive being jabbed with a set of nested electrodes and then having one pulled out. "All of those things, it seemed to me, made the possibility of this working less likely." Nonetheless, he told Jonas she could spend one afternoon a week working on this seemingly dead-end venture.

"So Friday afternoons, I started to put this thing together," says Jonas. "I had to learn to make electrodes. I had to figure out how I was going to move the electrodes past each other. I had to learn how to, by hand, put one electrode inside the other. Obviously I had a lot of problems to solve and I wasn't getting very far with one afternoon a week."

So she suggested to Kaczmarek that he allow her to pursue the project full time for one month. If she got the system to work, she would present the data at the upcoming biophysical society meeting, whose abstract was due in a month. "I guess I relented at that point," says Kaczmarek, "and she got it to work," two days before the abstract was due.

MITOCHONDRIAL MEMBRANE MYSTERIES

Jonas never did catch her white whale - the calcium channels that she thought would be embedded in vesicle membranes - although she did spot some IP3 receptor channels along the way. Instead, 15 years after her first summer with Llinás, Jonas and Kaczmarek returned to her old stomping grounds in Woods Hole to record the activity of what turned out to be channels in the membranes of mitochondria. They published the work in Science in 1999.

Those early days were very stimulating, says Jessica Helm, who worked with Jonas for three years before becoming a graduate student at the University of Stony Brook. "It seemed like every day there was something new to look at. When someone would find a channel, everyone would rush over. There were new ideas and new techniques and lots of diagrams drawn on napkins during coffee and cookies in the cafeteria. It was just really exciting for everyone involved."

It was also impressive to watch. "I remember when I met Liz, [I thought] that she was great. She was this wonderful woman doing such cool things," says Julie Kauer of Brown University, Kaczmarek's former student. "No one had ever done what she was doing before. Ever. In the universe. It just blew my mind."

Kauer says it also demonstrated that Jonas is fearless. "Who would ever think that such a crazy idea would work?" she says. "There's a reason no one had done it before. Maybe no one thought it could be done." Indeed, no one besides Jonas does it today. "I've talked to many other labs who have contemplated using this technique, but I don't know of anyone who's done it," says Marie Hardwick of Johns Hopkins University. "Usually they just collaborate with Liz."

As does Hardwick. Together, she and Jonas have found that proteins known to be involved in regulating cell death form channels in the mitochondrial membranes of living cells, where they influence neuronal signaling, thus governing whether a synapse will be strengthened or removed. "It's a marriage of two completely different fields," says Hardwick.

That marriage has not been all honeymoon. When the researchers started submitting papers together in 2002, Hardwick says, "we would get critiques from reviewers saying, ?This is a very strange model system, the squid.' Of course, scientists have won Nobel prizes studying squid, but it just seemed odd to investigators in the cell-death field who were not familiar with that area of research."

Jonas is now moving into mammalian systems, exploring the role that mitochondria and cell-death proteins might play in shoring up synapses during learning and memory. Although she hasn't yet been able to get her double-barrel electrodes to pick up mitochondrial channels in mouse hippocampal neurons, she and Hardwick have discovered that overexpressing Bcl-XL (a protein that staves off cell death) changes the structure of synapses: The modified neurons make more synapses, containing more synaptic vesicles, and their transmission is faster and better than control cells.

Jonas says she thinks that Bcl-XL and its relatives might enhance synaptic transmission by allowing mitochondria to release extra ATP. Given that releasing neurotransmitters is an energy-consuming business, the theory is not unreasonable. It is, however, controversial, because Jonas and her collaborators have found that the mitochondria are making more ATP while consuming less oxygen. "We think Bcl-XL is making the machine more efficient," says Jonas. "That's really controversial right now and we still have a lot of experiments to do," which doesn't bother her. "I've been on the frontier a lot," she says. "Whether I've done well with that is not clear."

khopkin@the-scientist.com