Cosmic rays represent a permanent factor involved both in biological evolution and in cellular ageing ( 1 ). During natural selection, organisms have developed mechanisms of biological/ biochemical defence against damages caused by space radiation in the earth's atmosphere.
On the basis of the studies performed in space, where cosmic rays are greater than those on the earth's surface, it was possible to highlight the main cellular targets and the possible consequences caused by cosmic radiation. Space experiments have demonstrated the formation of DNA strand breaks, induced mutations ( 2-4 ), lipidic peroxidation ( 5 ), cytokine secretion ( 6 ), but also an inhibition of proliferation, cellular differentiation, cell-cell interaction, and cytotoxic activity mediated by the immune system ( 2,6 ). Moreover, it has been observed that cosmic rays induce p53 expression, a protein that, causing apoptosis or blocking G1/S transition of cell cycle, safeguards genomic stability and protects against tumour formation induced by chemical and physical agents.( 2,3,7 )
It is interesting that different organisms and cells exposed to a small dose of radiation, or chemical mutagenic agents can become resistant to subsequent radiation dose exposures (adaptive response) ( 8 ). This cell adaptive response to cosmic rays exposure has been recently studied at the National Laboratories of Gran Sasso (LNGS) ( 9,10 ). LNGS are provided of an underground laboratory that gives the opportunity to evaluate the effects of a low radioactivity environment on various organisms. Biological experiments performed in the LNGS (Pulex and Pulex-2 projects) on yeasts ( 9 ) and on chinese hamster fibroblasts ( 10 ) suggest that the background radiation has an important role in determining various cellular adaptive processes. In particular, these experiments show that cells, grown at a low dose of radiation: 1) are less protected against DNA damages induced by chemical and physical agents; 2) are more prone to apoptosis; 3) have a different reaction in presence of oxidant agents.
Generally, on the one hand the cosmic rays influence the cell ageing and on the other hand they promote enzymatic systems able to protect the cells against radiation damages. It is likely that, while cosmic radiation influence positively the induction of defensive systems against radiation and other mutagenic agents on cells in culture, the low background radiation environment at the LNGS underground laboratory would not negatively affect the enzymatic systems of cryo-preserved cells. On the other hand, a reduced background radiation could favour maintenance of frozen cells, and thus their vitality and cell cycle restart (related with DNA strand breaks caused by cosmic rays) after thawing. These studies will be developed at the Research Center in Urbino (Centro di Citometria e Citomorfologia) that is a reference centre at an international level for the flow cytometric evaluation of the cell mortality and for the analysis of the haematopoietic stem cells (see pubblications 17-30 ).
On the base of the current knowledge of the biological effects of low doses of cosmic ray radiation on the activity of isolated cells ( 9,10 ), we plan to undertake the same project with a study of the same effects on animals, as complex organisms with an autonomous life.
From the numerous animal taxa which may be suitable models for this study, two phyla of aquatic microinvertebrates, Rotifera and Gastrotricha, seem to be more fitted to this purpose due to their biological and ecological characteristics ( 11,12 ). They are totally or partially eutelic (no mitotic divisions after hatching), have a short life cycles (lasting only weeks), and are able to produce some dormant stages ( 13 ), known to be more resistant against environmental stresses than the active forms ( 14 ). Dormant stages are easily induced under lab conditions by subtracting water, because these animals are naturally capable of resisting desiccation of their habitat ( 15,19,20 ). Bdelloid rotifers are peculiar for entering anhydrobiosis (= life without water) during any phase of their biological cycle: they desiccate their tissues severely and remain in a dormant but living stage till successive rehydration. That process has no cost on their life cycle, unless they meet unfavourable conditions during the dehydration, or a long duration of the dormancy period, or else accidents during the same phase, causing all these factors increased mortality ( 16 ). Tests in conditions protected from radiations, like those present in the underground laboratory, are planned for a comparison with similar tests carried out in an external laboratory. The experiments will be performed on 1) embryonic stages, 2) active forms and resistant forms.
The work plan on the microinvertebrates will be developed at the Center of Advanced Structural Analyses (C.A.S.A.) and at the Laboratory of Zoology of the Institute of Morphological Sciences of the University of Urbino, at the Department of Biology, Zoology Nat. Sc. section of the State University of Milan, where researchers have been dealing with experimental aspects of the biology of aquatic invertebrates for many years (see publications 11,12,15,16, 17, 18, 19,20 in the list below).
