Abstract
Golgi staining allows for the analysis of neuronal arborisations and connections and is considered a powerful tool in basic and clinical neuroscience. The fundamental rules for improving neuronal staining using the Golgi-Cox method are not fully understood; both intrinsic and extrinsic factors may control the staining process. Therefore, various conditions were tested to improve the Golgi-Cox protocol for vibratome-cut rat brain sections. Optimal staining of cortical neurons was achieved after 72 h of impregnation. Well-stained neurons in both cortical and subcortical structures were observed after 96 h of impregnation. The dendritic arborisation pattern of cortical neurons derived from the 72-h impregnation group was comparable to those of the 96 and 168-h impregnation groups. The entire brain was stained well when the pH of the Golgi-Cox solution was 6.5 and that of the sodium carbonate solution was 11.2. Lack of brain perfusion or perfusion with 0.9% NaCl did not influence optimal neuronal staining. Perfusion with 37% formaldehyde, followed by impregnation, only resulted in glial staining, but perfusion with 4% formaldehyde facilitated both glial and neuronal staining. Whole brains required longer impregnation times for better staining. Although every factor had a role in determining optimal neuronal staining, impregnation time and the pH of staining solutions were key factors among them. This modified Golgi-Cox protocol provides a simple and economical procedure to stain both neurons and glia separately.
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Acknowledgements
The authors gratefully acknowledge the constant support, encouragement and guidance from Dr. S. Gurumadhva Rao, President, RAKMHSU. The authors would like to thank the administration of United Arab Emirates University (UAEU), Al Ain, for providing research animals to carry out this work. Discussions with Dr. Tarig Hakim Merghani Hakim, Professor and Chairperson, Department of Physiology, RAKCOMS, RAKMHSU, have been insightful and we are thankful for the warm encouragement and support. The authors acknowledge the unconditional support and help from Dr. Kumar M. R. Bhat, Professor and Chairperson, Department of Anatomy, RAKCOMS, RAKMHSU. Help received from Ms. Sheela Haridas, Mr. Michael V. Magaogao, Mr. Nizam Arshad Khan, and Mr. Ramesh, Sr. Lab Technicians, RAKCOMS, RAKMHSU is gratefully acknowledged. The authors are indebted to the RAKMHSU for the financial support to carry out this research work.
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Narayanan, S.N., Bairy, L.K. & Srinivasamurthy, S.K. Determining factors for optimal neuronal and glial Golgi-Cox staining. Histochem Cell Biol 154, 431–448 (2020). https://doi.org/10.1007/s00418-020-01891-9
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DOI: https://doi.org/10.1007/s00418-020-01891-9