Phosphenes in space: a study on the interaction between carbon ions and rod photoreceptor

Phosphenes, or light flashes, have been reported by astronauts since the first Apollo flights to the Moon. They are usually described as occurring in the dark and typically before falling asleep. The light flashes are thought to originate as an effect of high- energy particles interacting with the visual system. The study presented in this thesis investigates the hypothesis of a direct excitation of the rod photoreceptors inside the retina and in particular the interaction between carbon ions and rods. Also the retinal and cortical electrophysiological response of mice submitted to carbon ions irradiation have been studied. This thesis has been developed as a section of the ’Anomalous Long Term Effects on Astronauts’ (ALTEA) program, and all the experiments concerning this thesis took place on ground. The objective of the ALTEA project is more general, and is aimed to assess the functional risks on the central nervous system due to particle flux in microgravity conditions during long term space mission. ALTEA includes a series of experiments both in orbit and on ground. The experiments on board International Space Station will contribute to define the causes of the anomalous phosphene perception of the astronauts by detecting the incident particles through the head of the astronauts and concurrently monitoring the central nervous system functionality. The experiments on ground include the direct irradiation with carbon ions of the outer segments of the rods, and an animal model to investigate the retina/cortex responses of mice irradiated in controlled laboratory experiments. In these experiments on ground I directly contributed in the preparation, the test and the optimization of the experiments and in the following analysis of the data. The whole project has been supported by the Italian Space Agency. The outline of this thesis includes an introduction to the interdisciplinary ALTEA program in the first chapter. In the second chapter ALTEA-MICE (Mice intermittent Irradiation with Concur- rent Electrophysiological monitoring) study is described: the experimental sessions as well as the evidences in the electrophysiological responses of the mouse retina have been described in detail. My role has been chiefly the experimental set up, and I also collaborated on the data analysis. The main results of this investigations have been published. In the third chapter the system of the rod outer segments in suspension and the mechanism of the activation of the photoreceptors are described. The fourth chapter deals with the radiation effects in the biological tissue, and in the fifth chapter there is particular regard to the effects of radiation on samples containing a suspension of rods extracted from bovine retinae. The effects on the rod photoreceptor rhodopsin has been studied: measurements have been performed irradiating with carbon ions the outer segments of the rods in suspension. The amount of the isomeric transformations (bleaching) of the molecules of chromophore into the photoreceptor gives the extent of the effects of radiation. I collaborated to the experiment and carried out the following data analysis. Finally, in the sixth chapter the process of the photo-isomerization of rhodopsin is explained by a model: the process starts with the water radiolysis and the formation of hydroxyl radicals OH in order to achieve the lipid peroxidation, then there is the subsequent emission of visible photons which are responsible for the photoreceptor bleaching. Since rhodopsin is surrounded by phospholipids, present in high concentration in the disc membranes, the effect of chemiluminescence is proposed to be the cause of the photo-transduction cascade and the light flash perception as a consequence. In order to test this hypothesis, a series of experiments was aimed at studying the effects of the hydroxyl radicals produced in an enzymatic way. The main results of this work are illustrated in chapter six, and have been published.

I fosfeni, o light flash, sono stati percepiti per la prima volta durante il volo verso la luna della navicella Apollo. In genere sono avvertiti al buio, prima di addormentarsi. In questa tesi si studia l'ipotesi di una eccitazione dei fotorecettori della retina da parte di ioni pesanti (carbonio). Campioni di recettori dei bastoncelli vengono irradiati, e viene studiato l'effetto degli ioni carbonio su topo, attraverso la registrazione della risposta elettrofisiologica (segnale retinico e corticale). La tesi si e' sviluppata nell'ambito del progetto ALTEA (Anomaluos Long Term Effect on Astronauts) e tutti gli esperimenti relativi a questo lavoro sono stati svolti a terra. L’obiettivo di ALTEA e’ piu’ generale e riguarda lo studio del rischio funzionale sul sistema nervoso centrale degli astronauti dovuto alla particelle ionizzanti presenti in condizioni di microgravita’ durante le missioni spaziali a lungo termine; ALTEA include sia esperimenti a terra che in orbita, a bordo della Sazione Spaziale Internazionale. La tesi inizia con una introduzione al progetto ALTEA; nel secondo capitolo e’ descritto lo studio chiamato ALTEA-MICE (Mice intermittent Irradiation with Concurrent Electrophysiological monitoring). Nel terzo capitolo sono descritto il sistema costituito dai segmenti esterni dei bastoncelli bovini in sospensione e il meccanismo di attivazione dei fotorecettori. Nel quarto capitolo e quinto capitolo mi sono occupata della interazione radiazione – materia nei tessuti e del caso specifico dell’irradiazione dei campioni dei segmenti esterni contenenti la rodopsina. Nel quinto e ultimo capitolo viene presentato un modello che spiega l’isomerizzazione (attivazione) del retinale della rodopsina come prodotto dell’azione dei radicali liberi provenienti dalla radiolisi dell’acqua.