## 4 Discussion
### Ca\({}^{2+}\), Endoplasmic Reticulum (ER), Mitochondria and Peroxisomes  
Ca\({}^{2+}\) is an important second messenger and plays a critical role in the fate of the cell surviving or dying. Currently, we have an observed increase in Ca\({}^{2+}\) with confocal microscopy during wound healing (24 h post amputation) but a decrease in day 5 blastema. It is possible that increased Ca\({}^{2+}\) content derived from intracellular Ca\({}^{2+}\) stores, such as ER, following plasma exposure might be responsible for apoptosis of damaged cells during the wound healing period. Low Ca\({}^{2+}\) in the blastema of experimental tadpoles may support early differentiation of these cells as reported in mammalian epidermal cells [77], resulting in faster tail regeneration in experimental tadpoles.  
Additionally, the relationship between Ca\({}^{2+}\) and ROS is complicated and close, either as a crucial partner in regulating the redox status of cells, determining cell fate, or in signaling in response to a number of physiological and stress conditions [78]. Our studies find support from the work of [PERSON] et al. [29] that an increase in intracellular ROS [33] and Ca\({}^{2+}\) following APP exposure (current study) could serve as valuable biomarkers for the oxidative stress.  
Mitochondria also undergo remodeling of their shape and morphology during programmed cell death--a biological process that usually confers an advantage during an organism's life-cycle. Mitochondria are dynamic organelles that continually undergo fusion and fission. These are two opposing processes that act in concert to maintain the shape, size, and number of mitochondria as well as their physiological function [79]. Our electron microscopy study shows that some mitochondria from experimental tadpoles had fused acquiring elongated or giant shapes with wide cristae compared to controls where mitochondria were round, smaller and with thin cristae. A high level of stress leads to fission of mitochondria, which would eliminate damaged mitochondria through mitophagy [80]. Mitochondrial fission also inhibits propagation of the Ca\({}^{2+}\) signal and protects against Ca\({}^{2+}\) mediated cell death [81], whereas mitochondrial fusion allows them to compensate for one another's defects by sharing components, in order to maintain energy output in stressful situations [82; 83]. Fission and fusion interact to create a diversity of mitochondrial structures and ER may play an active role in determining the sites of mitochondrial fission [84]. Additionally, in experimental tadpoles, some mitochondria were swollen with an electro-lucent matrix compared to control, which could be due to ischemia [85], increased Ca\({}^{2+}\) overload [18; 55], and induction of mPTP [86]. Increase in Ca\({}^{2+}\) (as observed using the confocal microscopy), as well as oxidative stress [33] in the regenerated tissue following plasma exposure, might be responsible for the opening of mPTP, which changes mitochondrial morphology as reported by [PERSON] and [PERSON] [87]. Currently observed changes in mitochondrial dynamics may allow cells to adapt to oxidative stress which warrants further studies.  
ER is also known to generate peroxisomes [88]. Currently, an observed increase in peroxisomes in blastema of experimental tadpoles might be generating and releasing important signal molecules; such as, O\({}_{2}^{-}\), H\({}_{2}\)O\({}_{2}\) and NO in cytosol, which can contribute to a more integrated communication system among cell compartments as reported by [PERSON] et al. [89]. Additionally, peroxisomes must be providing protective mechanisms to counteract oxidative stress.