Deborah Nelson, Ph.D.

Chloride channels and synaptic function.

Synaptic development and plasticity are integral to our understanding of neuronal function and disease, so understanding the events that underlie these processes, including the role of ClC-3 chloride channels, is of fundamental significance. Chloride channels are a physiologically important, yet under-studied, class of channels in the brain. A role for depolarizing GABA-mediated Cl- driven excitation is now widely accepted as a model for neuronal maturation in cortical circuits; however, a role for ClC channels in the phenomenon has not previously been explored. ClC-3 is unique among its family members in that it is gated by Ca2+ dependent phosphorylation. Our recent published studies 39, demonstrate that ClC-3 channels colocalize with NMDA receptors at hippocampal synapses. Calcium flowing through NMDA receptors activates ClC-3 channels enhancing the excitatory postsynaptic potential (EPSP) in immature neurons and reducing the EPSP in mature neurons. Long-term potentiation (LTP) and its dependence upon NMDA-receptor mediated calcium entry has been widely studied as a mechanism underlying learning and memory. Based upon our data, expression of ClC-3 channels would enhance NMDA-receptor dependent calcium signaling and thereby facilitate the induction of long-term potentiation at early times in development and conversely depress both calcium signaling and induction of LTP in mature neurons. Thus, Ca2+-dependent ClC-3 channels and their unique relationship with NMDA receptor provide a new and important level of regulation in the modulation of synaptic plasticity. In that ClC-3 channels are ubiquitously expressed in the brain, the relationship between ClC-3 channels and NMDA receptors is not likely to be restricted to the hippocampus, however, this remains an open question. Our current studies are targeted at an integrated understanding of the role postsynaptically expressed ClC-3 plays in the fundamental aspects of hippocampal neuronal excitability as well as synaptic plasticity.

Chloride channels in pancreatic beta cell function

The most prevalent form of diabetes (type II; 90% of cases) is characterized by insulin resistance in peripheral tissues and/or a deficiency of insulin due to the failure of pancreatic beta-cells to secrete insulin. Considerable progress has been made in determining many features of secretagogue-regulated insulin secretion, particularly the role of plasma membrane ion channels in the process. However, we are still at an early stage in defining the molecular mechanisms involved in exocytosis of the insulin granule, and the possible role of granule membrane ion channels in this process. Previously, we showed that the chloride channel ClC-3 is functionally expressed in the membrane of insulin-containing granules. Functional studies in isolated beta-cells showed that activation of ClC-3 is permissive for insulin secretion. This is due, at least in part, to the promotion of granular acidification; various strategies to abolish acidification disrupt secretion in a similar manner. These observations are part of a burgeoning literature on the important role played by vesicular ion channels in secretion, as well as the more specific requirement for vesicle acidification in several cases. Recently, we have extended our findings to the ClC-3 knockout mouse. Our recent data indicate that beta-cells from this model are defective in exocytosis and animals exhibit aberrant glucose tolerance, indicating that they may become diabetic. Current investigations in the laboratory build on this foundation and are directed at unraveling in molecular detail the role of ClC-3 chloride channels in beta-cell secretion.

Ion channels in macrophage function

Several years ago our studies started out as purely cell physiological in design and implementation. These studies have encompassed ion channel function, phagocytosis and organellar acidification, principally in macrophages. Historically, it is well known that basic studies inform etiology and therapeutics in various diseases, thus we are obliged to consider the clinical implications of our own findings. Patients with CF are highly susceptible to chronic bacterial infection, with the lung being the most clinically important site. Mutations in CFTR lead to an increased propensity for pulmonary infection in susceptible subjects and morbidity is usually associated with lung damage. Despite extensive knowledge of mutations in the Cftr gene as well as the biophysics and cell biology of the chloride channel that it encodes, the connection between fatal lung infection and defective CFTR channel function is unclear. The mechanism by which the genetic defect in CF leads to bacterial respiratory infections and associated chronic inflammation remains one of the major obstacles to the design of long-term effective therapeutic strategies. While compromised epithelial cell function and resultant changes in the mucus layer are important factors in promoting bacterial persistence, the role of the innate immune system is still uncertain. It is well-established that neutrophil-dependent airway inflammation exists in CF but we propose that additional deficiencies may be attributable to a failure of alveolar macrophages and possibly neutrophils to exhibit vigorous bactericidal activity. Defects in the behavior of the innate immune system could thus have important consequences for microbial defense in CF patients, and we feel that this is a potential therapeutic avenue that should be explored further. It
should be borne in mind, however, that a continued emphasis remains on the basic relationship between ion channel function and organellar acidification in these cells. We hold that further understanding of this issue has wide implications for organelle biology as well as expanding our knowledge of ion channel regulation.