Relative Resilience of Cerebellar Purkinje Cells in a Cardiac Arrest/Resuscitation Rat Model

Abstract

Background

In studies on cardiac arrest (CA)/resuscitation (R) injury, Purkinje cell degeneration was described, however, with inconsistent data concerning severity and time point of manifestation. Moreover, CA/R studies paid only limited attention to inhibitory stellate interneurons. To this aim, the hypothesis that cerebellar could be relatively resilient toward CA/R because of diverse cellular defense mechanisms including interaction with stellate cells was tested.

Methods

We examined rats with survival times of 6, 24, and 48 h, and 7 and 21 days in comparison with sham- and nonoperated animals. Thereby, we focused on the immunohistochemical expression of cfos, MnSOD, Bcl2, caspase 3, parvalbumin, calbindin D28 k, MAP2, IBA1, and GFAP, especially in the particular sensitivity to CA/R cerebellar lobule IX. Hippocampal CA1 degeneration was demonstrated by expression patterns of MAP2 and NeuN in combination with IBA1 and GFAP.

Results/Conclusions

Comparative analysis of hippocampal CA1 pyramidal cells and cerebellar Purkinje cells confirmed a relative resil-ience of Purkinje cells to CA/R. We found only a notable degeneration of Purkinje cell neuronal fiber network, which, however, not necessarily led to neuronal cell death. To induce significant Purkinje cell loss, a stronger ischemic trigger seems to be needed. As possible Purkinje cell-protecting mechanisms, we would propose: (1) activation of inhibitory stellate cells, shown by cfos, MnSOD, and Bcl2 expression, balancing out ischemia-induced excitation and inhibition of Purkinje cells; (2) translocation of the calcium-buffering system, shown by parvalbumin and calbindin D28 k expression, protecting Purkinje cells from detrimental calcium overload; (3) activation of the neuron–astrocyte cross talk, protecting Purkinje cells from over-excitation by removing potassium and neurotransmitters from the extracellular space; (4) activation of the effective and long-lasting MnSOD defense system; and (5) of the anti-apoptotic protein Bcl2 in Purkinje cells itself. Moreover, the results emphasize the limited comparability of animal CA/R studies because of the heterogeneity of the used experimental regimes.

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Abbreviations

Bcl2:

B cell lymphoma 2

CA/R:

Cardiac arrest/resuscitation

DAPI:

6-Diamidino-2-phenylindole

GCL:

Granule cell layer

GFAP:

Glial fibrillary acidic protein

HNF4A:

Hepatic nuclear receptor 4A

IBA1:

Ionized calcium-binding adaptor molecule 1

ID:

Integrated density

IPPV:

Intermittent positive pressure ventilation

LD:

Linear density

MAP2:

Microtubule-associated protein 2

ML:

Molecular layer

MnSOD:

Manganese-dependent superoxide dismutase (mitochondrial SOD 2)

NeuN:

Neuronal nuclei antibody

PBS:

Phosphate-buffered saline

PC:

Purkinje cell

PCL:

Purkinje cell layer

PFA:

Phosphate-buffered paraformaldehyde

ROSC:

Return of spontaneous circulation

ST:

Survival time

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Acknowledgements

The authors wish to thank Leona Bück for the excellent technical assistance.

Funding

None.

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Authors

Contributions

All authors have made substantive contributions to the study. GK conceived the study, was responsible for data analyses, writing the manuscript and preparing the illustration of results. TMNT carried out all immunohistochemical stainings. TE carried out surgery including post resuscitation care and tissue sampling. UE established the experimental model and supervised the animal experiments and critically reviewed the manuscript.

Corresponding author

Correspondence to Gerburg Keilhoff.

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The authors declare no conflict of interest.

Ethical Approval

This study was granted according to the requirements of the German Animal Welfare Act on the Use of Experimental Animals and the Animal Care and Use Committees of Saxony-Anhalt (permit number 42502-2-2-947 Uni MD).

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Keilhoff, G., Nguyen Thi, T.M., Esser, T. et al. Relative Resilience of Cerebellar Purkinje Cells in a Cardiac Arrest/Resuscitation Rat Model. Neurocrit Care 32, 775–789 (2020). https://doi.org/10.1007/s12028-019-00799-0

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Keywords

  • Bcl2
  • Calcium-binding proteins
  • Caspase 3
  • cfos
  • MnSOD
  • Stellate cells