## 3 The Effect of RF Exposure on _in Vitro_ Models: Finding the Pathways
### The Effect of RF on Neuronal Cell Activity  
The interest in RF exposure effects on the central nervous system has grown in parallel with mass technology applications. Given the proximity to the area involved in mobile phones handling, several studies analyze cells from facial districts such as eyes and nose, which are doors to the nervous system. In particular, no effect of exposure to a very high SAR (20 W/kg) was observed when retinal ganglion cells were exposed to three mobile phone frequencies (GSM-900, GSM-1800, UMTS). The eye represents an easy target for RF and the damage could compromise not just local tissue but also general visual CNS function. However electrophysiological evidence from single cell recording could not determine any frequency, or intensity or distance-related damage after RF exposure [36]. Similarly, no change was detected in physiological parameters in the nasal area [37] or in the oral mucosa cells both spatially close to cell phone EMF exposure [38]. Regarding neuronal networking, [PERSON] [39] published a report about the thermal effect of short-term exposure to GSM-1800 on neuronal firing by measuring the spontaneous electrical activity _in vitro_. In this case, the increase of firing activity detected was ascribed to the increase of localized temperature that might influence general EEG and neuronal tissue functions _in vivo_. On the other hand, 1 C increase in temperature after EMF exposure did not elicit activation of heat shock protein in human brain cells [40].  
One of the most frequent findings of _in vitro_ models after EMF-RF exposure is the increase of oxidative stress-related events that lead to cell damage. This event is causally related to neurodegeneration that underlie severe pathologies such as Alzheimer's disease or Parkinson's. [PERSON] [41] investigated the stress-related effect of RF-EMF pulse exposure on dopaminergic MN9D cultures. The increase of intracellular nitric oxygen (NO) level after exposure was related to the activation of Ca/CaM-dependent constitutive nitric oxide synthase. The CaM/NO/cGMP signaling pathway activates a rapid response cascade that in turn influences EMF on Cam/NO pathways, which can modulate CNS response to inflammation and ischemia and interfere with the ability of the brain to restore post-traumatic functions. Another mechanism to regulate neuronal function is calcium homeostasis, suggested to be a mediator of cell response to RF exposure [42]. While [PERSON] [43] found no effect on calcium homeostasis in cultured hippocampal neurons or peripheral human endothelial cell exposed to 900 MHz GSM-modulated RF fields (SARs of 2 W/kg to meet guidelines set by ICNIRP), a growing bulk of literature indicates calcium-related machinery as a potential mediator for EMF effect, both _in vitro_ and _in vivo_. [PERSON] [44,45] hypothesized that repeated five-hour sessions of daily exposure to rats of 835 MHz (SAR = 1.6 W/kg) might alter permeability of cell membrane in hippocampal pyramidal cells, thereby jeopardizing neuronal connectivity.  
Regarding the peripheral system, oxidative stress is a critical factor that also interferes with immune system cell viability. Peripheral blood mononuclear cells are a critical component of the immune system that fight infection and adapt to intruders and are also involved in aging and CNS neurodegeneration. EMF-RF induces lipid and protein expression changes leading to reactive oxygen species (ROS) production and caspase-3-dependent apoptosis through weakening mitochondrial membrane potential [46].  
What emerges from these reports, and several earlier ones, is that RF exposure is bad for cell survival and that there are some districts such as the brain, in which cells are extremely sensitive to RF exposure. However, we would like to point out a few examples that might suggest otherwise.