Abstract
Heterologous channel expression can be used to control activity in select neuronal populations, thus expanding the tools available to modern neuroscience. However, the secondary effects of exogenous channel expression are often left unexplored. We expressed two transient receptor potential (TRP) channel family members, TRPV1 and TRPM8, in cultured hippocampal neurons. We compared functional expression levels and secondary effects of channel expression and activation on neuronal survival and signaling. We found that activation of both channels with appropriate agonist caused large depolarizing currents in voltage-clamped hippocampal neurons, exceeding the amplitude responses to a calibrating 30 mM KCl stimulation. Both TRPV1 and TRPM8 currents were reduced but not eliminated by 4 hr incubation in saturating agonist concentration. In the case of TRPV1, but not TRPM8, prolonged agonist exposure caused strong calcium-dependent toxicity. In addition, TRPV1 expression depressed synaptic transmission dramatically without overt signs of toxicity, possibly due to low-level TRPV1 activation in the absence of exogenous agonist application. Despite evidence of expression at presynaptic sites, in addition to somatodendritic sites, TRPM8 expression alone exhibited no effects on synaptic transmission. Therefore, by a number of criteria, TRPM8 proved the superior choice for control over neuronal membrane potential. This study also highlights the need to explore potential secondary effects of long-term expression and activation of heterologously introduced channels.
Highlights
There has been strong recent interest in heterologous control over electrical activity in neurons and other excitable cells [1,2,3]
Our observations suggest that transfection of transient receptor potential vanilloid receptor 1 (TRPV1) alone, in the absence of capsaicin stimulation, has important effects on neuronal function
We did not detect overt cell loss or swelling of capsaicin-naıve TRPV1-transfected cells in estimates of transfection efficiency 24 hr following transfection, cell-attached patch recordings from TRPV1-transfected cells exhibited markedly more seal instability (Figure 1C) and spontaneous rupture compared with TRPM8-transfected cells (Figure 3C)
Summary
There has been strong recent interest in heterologous control over electrical activity in neurons and other excitable cells [1,2,3]. These approaches offer the possibility of remotely controlling the activity of select populations of neurons, thereby gaining experimental or therapeutic influence over network activity. Introduction of leak channels [17,21,22] suffers from a lack of control over the degree of activity change and typically is limited to an inhibitory influence, and one obvious disadvantage of optogenetic approaches is the need for a light source. Some applications may require activation of a spatially dispersed set of neurons over a large area, in which case a single light source may not be adequate
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