**3.2 Pathogenic studies and mechanisms in favor of neurotropic nature of the SARS CoV-2**

Reports associated with neurological impairments induced by CoV-2 in acute as well as post-acute infection stages have been accumulated since early 2020 [71, 72]. Whether these effects are occurring as a result of the neuroinvasive nature of the virus or due to the overt immune responses is not yet fully understood. A recent study in medRxiv reveals that in comparison to increased inflammatory and cytokine storm markers found in the serum of COVID-19 patients, their levels are rather low in the CSF. This was corroborated by comparing and contrasting the insignificant neuroinflammatory changes in these patients' CSF compared with CSF of patients with autoimmune pathologies that displayed very high neuroinflammation. On the contrary, a significant increase in CSF Neurofilament-L (NF-L) in critical cases suggests neuroaxonal injury and strengthens the neurotropic nature of the virus [73]. The olfactory transmucosal pathway has been an established port of entry for CoV-2 virus, but we only have a limited knowledge about the virus-host interactions [3]. The human sequencing data point toward the role of supporting cells of OE in the viral entry. This is because of the expression of ACE2 and TMPRSS2 proteins in these cells that serve as the entry factors [5]. The immunohistochemical analysis of the SARS-CoV-2 S protein, however, revealed a characteristic granular, perinuclear expression pattern in olfactory mucosal cell types, which were of neuronal origin (revealed by expression of Tuj1, Neurofilament 200, and Olfactory Marker Protein) obtained from the autopsy samples of the COVID-19 infected patients [3]. Additionally, presence of CoV-2 particles was confirmed in the CNS regions including the OB [74]. This questions the current understanding of non-neuronal vs. neuronal viral infection occurring in COVID-19. Generally, neurotropic viruses access the peripheral regions to gain entry into the CNS [75]. Whether the CoV-2 is causing neuronal pathogenesis directly or indirectly is not fully understood. We also do not have a complete understanding of the virus' pathway to the OB and other CNS regions. In an attempt to investigate the pathogenic mechanisms of the virus, an in vitro study of generating human sensory neurons from the human embryonic stem cell lines was carried out. These peripheral sensory neurons were shown to express ACE2 and were indeed receptive to the virus, which is in contrast with the reported non-neuronal expression of ACE2. One hour after the incubation with the virus, intracellular expression of nsp-14, S protein, RdRp, and nucleocapsid phosphoprotein viral genes was substantially upregulated in the infected neurons [76]. The molecular pathologies relating to chemosensory perception were specifically affected in the infected peripheral neurons. The human induced pluripotent stem cells (hIPSCs)-derived midbrain dopaminergic neurons were shown to be selectively permissible to the CoV-2 infection. Further, inflammatory and cellular senescence responses were observed in these neurons both in vitro and upon transplantation in vivo as well [77]. In another attempt to investigate if human neurons are a direct target of this virus, three-dimensional human brain organoids system was utilized. Preferred tropism to mature neurons of the cortical plate in relatively older brain organoids (day 60) was found out. The virus has relatively lesser influence on the actively proliferating neural precursor cells of the ventricular zone of young (day 15) organoids. Moreover, CoV-2-infected neurons displayed mislocalized Tau protein in their soma, which can potentially cause cellular stress reactivity and toxicity [78].
