However, maintenance of live colony is costly and sometimes difficult. Cryopreservation of germplasm circumvents the need for maintenance of live colony and genetic material would still be available for future use. In addition, up to now, many inbred mutant and genetically modified rat strains have not been readily available to investigators around the world [1], [28], [31] and [49]. Cryobanking of embryos, sperm, oocytes are becoming

very important both for reducing the maintenance cost and improving distribution of strains [1] and [36]. Cryopreservation of sperm provides a simpler and more economical alternative to cryopreservation of embryos, Ganetespib supplier and reduces the cost and space needed for keeping a large number of rat strains having a single mutation [1] and [35]. Sperm preservation protocols vary among species due to their inherent characteristics. There are marked species differences

in spermatozoa size and morphology. In addition, there are also more subtle differences in membrane phospholipid composition and metabolism of spermatozoa [6]. Rat sperm are known to have extreme sensitivity to suboptimal conditions such as centrifugation, pipetting, chilling, osmotic stress [34], [46] and [51] freezing and thawing [25], [34] and [35] possibly due to unusually long tail, head shape and membrane composition [12], [20] and [24]. Thus, acceptable and repeatable rat sperm cryopreservation protocol has not been achieved [57]. Post-thaw find more sperm quality is still unsatisfactory for intrauterine insemination Carnitine palmitoyltransferase II or in vitro fertilization in rats with genetic modifications [34] and [57]. Despite species variation, there are common stages to any sperm freezing protocol. All protocols involve sperm collection and extension, addition of cryoprotective agents (CPA) and cooling above 0 °C, freezing below 0 °C, storage and thawing [11]. During all of these stages, spermatozoa are exposed a number of potentially damaging stresses such as the change in temperature, osmotic and toxic stresses presented by exposure to high molar concentrations

of CPA and the formation and dissolution of ice crystals in the extracellular space [54]. Success of cryopreservation depends on sperm endurance to these insults [45] and [54]. Extenders, CPA, optimal cooling and thawing rates play important role for successful cryopreservation of sperm [10], [20], [30] and [42]. Extender composition and cooling rate have significant effects on sperm viability and there is a strong interaction between extender and cooling rate [55]. If the cooling rate is slower or faster than optimum cooling rate, this may cause irreversible damage to sperm [13], [27] and [29]. An optimum cooling rate must be slow enough to permit water to leave the cells to avoid intracellular ice formation, and fast enough to avoid severe cell dehydration and cryo-injury due to the solution effect [29].