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The Na+/H+ Exchanger and the Nigrostriatal Dopamine System

by
Marcelo de Avilez Rocha
BA
University of Maryland
Baltimore County

Thesis Advisor: Patricia Sonsalla, PhD

Graduate Program of Neuroscience

RWJMS Room V-10

Wednesday, April 30, 2008
3:00 pm


Abstract

Striatal dopamine (DA) neurotransmission facilitates motor behavior and is disrupted in patients afflicted with Parkinsonís disease (PD) due to the loss of nigrostriatal DA neurons. Although the causes of PD are unknown, pathogenic defects of mitochondrial respiration, metabolic stresses, reactive oxygen species, and disturbances in vesicular DA storage have been implicated. In the brain, Na +/H + exchanger (NHE) proteins are thought to regulate intracellular pH and volume of pre-synaptic axon terminals and may influence neurotransmission. Moreover, the NHE isoform 1 (NHE1) has been linked with neuronal damage during metabolic stress. Yet little is known of the role of NHE proteins in striatal DA neurotransmission or in the damage of nigrostriatal DA terminals caused by metabolic stress.

Based on existing knowledge of NHE proteins in other brain regions, it was hypothesized that NHE proteins (1) influence striatal DA neurotransmission, (2) participate in DA terminal damage caused by metabolic stress, or both. These hypotheses were tested pharmacologically using in vivo microdialysis in mice or in vitro using rat mesencephalic primary cell cultures. Expression of mRNA for different NHE isoforms was explored in nigrostriatal regions with RT-PCR. Localization studies for NHE1 protein were achieved with western blot analysis and double-label immunofluorescence confocal microscopy.

Microdialysis studies revealed that striatal NHE inhibition elicits a transient increase followed by a progressive reduction in striatal DA neurotransmission in mice. These changes were accompanied by an increase in DA turnover and a reversible decline in striatal tissue DA content, which are reminiscent of the effects typically obtained with inhibitors of vesicular DA storage. Striatal NHE inhibition reduced DA overflow caused by the mitochondrial inhibitor malonate, but it did not modify the malonate-induced damage to nigrostriatal DA terminals. Expression of mRNA for NHE1-5 was detected in the striatum and ventral midbrain. Moreover, protein localization studies revealed that NHE1 is expressed at the synaptic level, in close association but not directly co-located with nigrostriatal DA neurons. Studies in mesencephalic cell culture confirmed that NHE inhibition did not modify damage of DA neurons caused by various mitochondrial inhibitors.

These studies provide the first demonstration that striatal NHE inhibition modifies DA neurotransmission in vivo possibly via an effect on vesicular storage. The pharmacological effects are further supported by the detection of mRNA for NHE1-5 in the striatum and the location of NHE1 protein at the synaptic level. Given the absence of NHE1 immunoreactivity in nigrostriatal DA neurons, it is inferred that the observed effects on neurotransmission occur via inhibition of NHE1 located on non-DAergic cells or via inhibition of multiple NHE isoforms. Moreover, the data indicate that NHE1 is not involved in the damage of nigrostriatal DA neurons in the malonate model of metabolic stress. Together these data support a role for NHE proteins in striatal DA neurotransmission and raise new implications for the current understanding of nigrostriatal DA neuron homeostasis.


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