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. 2017 Jul 1;58(4):439-445.
doi: 10.1093/jrr/rrx004.

Effect of Acute Millimeter Wave Exposure on Dopamine Metabolism of NGF-treated PC12 Cells

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Free PMC article

Effect of Acute Millimeter Wave Exposure on Dopamine Metabolism of NGF-treated PC12 Cells

Alexis J Haas et al. J Radiat Res. .
Free PMC article

Abstract

Several forthcoming wireless telecommunication systems will use electromagnetic frequencies at millimeter waves (MMWs), and technologies developed around the 60-GHz band will soon know a widespread distribution. Free nerve endings within the skin have been suggested to be the targets of MMW therapy which has been used in the former Soviet Union. So far, no studies have assessed the impact of MMW exposure on neuronal metabolism. Here, we investigated the effects of a 24-h MMW exposure at 60.4 GHz, with an incident power density (IPD) of 5 mW/cm², on the dopaminergic turnover of NGF-treated PC12 cells. After MMW exposure, both intracellular and extracellular contents of dopamine (DA) and 3,4-dihydroxyphenylacetic acid (DOPAC) were studied using high performance liquid chromatography. Impact of exposure on the dopamine transporter (DAT) expression was also assessed by immunocytochemistry. We analyzed the dopamine turnover by assessing the ratio of DOPAC to DA, and measuring DOPAC accumulation in the medium. Neither dopamine turnover nor DAT protein expression level were impacted by MMW exposure. However, extracellular accumulation of DOPAC was found to be slightly increased, but not significantly. This result was related to the thermal effect, and overall, no evidence of non-thermal effects of MMW exposure were observed on dopamine metabolism.

Keywords: 60 GHz; dopamine turnover; in vitro; neuron-like model.

Figures

Fig. 1.
Fig. 1.
Experimental protocol. In order to assess the impact of MMWs on cell metabolism, while identifying any outcome unrelated to the thermal effect, PC12 cells in 6- or 96-well tissue-culture plates were exposed to three different sets of experimental conditions: an exposure to MMWs at 60 GHz, a heat increase in the medium induced by the thermostat setting of the incubator (heat control: HC) or a sham-exposure (MMW generator turned off).
Fig. 2.
Fig. 2.
Impact of MMWs exposure on intracellular DA contents in NGF-treated PC12 cells. (A) The metabolic phenotype of NS-1 cells was assessed by measuring the intracellular DA and NE contents after 24 h of NGF treatment (N = 4, mean ± SEM *P < 0,05). Statistical significance is represented by an asterisk. (B) Intracellular contents of DA and (C) intracellular contents of DOPAC, were determined by HPLC-ECD following a 24 h exposure to MMWs. The results are expressed as ng/106 cells (N = 5, mean ± SEM). (D) The DOPAC/DA ratio was also determined (Fig. 2D) (N = 7, mean of ratios from each experiment ± SEM).
Fig. 3.
Fig. 3.
Impact of MMWs exposure on extracellular DA and DOPAC contents in NGF-treated PC12 cell media. (A) DA and (B) DOPAC contents determined by HPLC-ECD in the medium of differentiated PC12 cell cultures after 24 h exposure to MMWs. The results are expressed in ng/ml of medium (N = 10, mean ± SEM). (C) Kinetics of DOPAC accumulation in the medium of NGF-treated PC12 cell cultures: DOPAC contents were measured 0, 10, 120, 480 and 1440 min (24 h) after initial medium change at t = 0 min (N = 4, mean + SEM). (D) DOPAC accumulation inhibition was assessed by adding 10 µM of the MAO inhibitor pargyline to the media (t = 0). DOPAC contents in the media were measured at t = 180 min (N = 4, mean ± SEM).
Fig. 4.
Fig. 4.
Impact of MMWs exposure on DAT protein expression in NGF-treated PC12 cells. (A) Microscopy images of DAPI (left panel) and DAT (right panel) fluorescent staining in NGF-treated cells: sham condition (top panel) vs MMWs exposure condition (bottom panel); scale bar = 10 µm. (B) Mean DAT and (C) mean housekeeping protein β-actin fluorescence intensity analyzed cell by cell. Data are expressed as fold of sham condition expression (N = 4, mean of means, ± SD *P < 0,05). Statistical significance found using one-way ANOVA is represented by an asterisk. (D) Distribution of DAT fluorescent staining intensity within NGF-treated cell populations (N = 4, pooling of the data from of all experiments).

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