Nerve cells can also release neurotransmitters outside of synapses

Every neurobiology textbook invariably states that nerve cells communicate with each other through synapses, the specialized cell-cell contacts found at the end of the cells' threadlike extensions. In the journal Science, researchers at the Salk Institute for Biological Sciences and the University of California at San Diego report that nerve cells, or neurons, may not have to rely on traditionally defined synapses to "talk" to each other.

The new study indicates that nerve cells can also release neurotransmitters outside of synapses. Neurotransmitters are the chemical messengers that nerve cells use to shuttle outgoing signals to adjacent neurons. Salk scientists refer to the release of neurotransmitters outside of synapses as "ectopic neurotransmission".

Cell-cell communications in other parts of the nervous system may also rely on ectopic neurotransmission, the new study suggests. This finding is an additional challenge to the descriptions of neuronal signal transmission found in neurobiology textbooks.

"Our results opened up the possibility that neurons can communicate many other ways not just at the traditional places that are defined by their anatomy," says lead investigator Terrence J. Sejnowski, who heads the Crick-Jacobs Center for Computational and Theoretical Biology at the Salk Institute. To investigate cell-to-cell communication in the nervous system, Sejnowski and his colleagues developed a computer model simulating signal transmission at a particular synapse in chick embryos. The computer model convinced Sejnowski and his collaborators that it may be time to rethink cell-to-cell communication in the nervous system.

In the past, the suggestion that neurotransmitters could be released and find their targets outside of clearly defined synapses, was considered an almost heretical notion.

A unique collaboration between anatomists and physiologists at the University of California in San Diego and theoretical neurobiologists at the Salk Institute was needed to rethink the standard model of neurotransmission. Darwin Berg in the Biology Department provided the physiological data upon which the model was based and Mark Ellisman in the Department of Neuroscience created a high-resolution reconstruction of the structure using high-voltage electron microscopy.

"In addition to the discovery of ectopic transmission this is the very first time that all these elements have been brought together," says Sejnowski. "Combining mathematical modeling with physiological, anatomical and behavioral data is the future of neurobiology. It allows us to draw conclusions that we could not have reached in any other way."

According to textbooks, nerve signals are transmitted from cell to cell only via specialized junctions. The transmitting neuron has a slightly swollen terminal point, which houses small vesicles that are filled with neurotransmitter. Upon arrival of a nerve signal, the vesicles spill their content into the narrow space between two cells. The released neurotransmitter molecules flow across the gap to the adjacent nerve cell and bind to specific receptor proteins on the receiving cell's membrane. If the receiving cell is a neuron, the binding of the neurotransmitter will generate an electrical impulse that travels along the length of the cell. If the receiving cell is a muscle cell, it will be stimulated to contract.

Based on high-resolution electron microscope images, research fellow Jay S. Coggan and scientist Thomas M. Bartol, the co-first authors of the Science paper, created a highly accurate computer model simulation of the giant chick embryo synapse that connects nerve fibers originating in the brain with the neuron that controls the size of the pupil and the shape of the eye lens. This particular synapse is a favorite model for the study of synaptic transmission since the neurons forming what is known as the ciliary ganglion are rather simple, easily accessible and nerve impulses can be recorded from either the presynaptic or the postsynaptic element or both.

Such recordings of nerve signals tipped off the researchers to the possibility that receptors outside of the synapse were routinely activated. But where did the necessary neurotransmitter molecules come from?

Coggan and Bartol then simulated the release of neurotransmitter from single vesicles located within synapses as well as from vesicles located outside traditional synaptic junctions in what they called "ectopic release". Next they compared their predictions with actual recordings from living cells. "We could only match the physiological results when we allowed 90 per cent of the release to occur outside of synapses," says Bartol who remarks in a philosophical aside: "Ectopic release is an interesting term because it means 'out of place' but of course in nature nothing is out of place."

"I think it challenges how we define synapses. There might still be synapses that fit the old definition but that might not be the only way that cells communicate," adds Coggan.

The necessary machinery, for one, has always been there. Scientists had known for a long time that all over the nervous system neurotransmitter-filled vesicles, SNARE-complexes responsible for fusing vesicles with cell membranes and neurotransmitter receptors were sitting outside of synapses but nobody knew what their function was. "A lot of the signals we recorded from the ganglion indicated that receptors that were considered perisynaptical were being activated in large numbers," remembers Coggan. "For me it was very gratifying when the computer model verified our suspicions."

Coggan points out that traditional and ectopic neurotransmission might simply serve different functions since acetylcholine, the neurotransmitter bridging the synaptic gap in the ciliary ganglion, activates two different types of nicotinic receptors: Alpha7-acetylcholine receptors (á7-AchR) and alpha3- acetylcholine receptors (á3-AchR) that differ in their biophysical properties and their spatial distribution. á3-AchRs cluster at synaptic sites on the cell body whereas á7-AchRs are found in perisynaptic areas. Ectopic transmission activates almost exclusively the latter.

"We are just beginning to appreciate the diversity and complexity of signaling in the brain. In the past we have been guided by very simple model systems like the neuromuscular junction. But in the brain there is an enormously more complex set of connections and demands that the system has to be able to perform. Evolution could have created entirely new possibilities," says Sejnowski.

The Crick-Jacobs Center for Computational and Theoretical Biology, funded by a generous gift from Joan and Irwin Jacobs, uses computer-based computational methods to mine the enormous amount of data on the composition of genes and proteins in the brain as well as the neural networks that regulate information processing. The ultimate goal is to generate theoretical models that explain how the brain works.