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Genetically Encoded Calcium Indicators Can Impair Dendrite Growth of Cortical Neurons
Ina Gasterstädt1, Alexander Jack1, Tobias Stahlhut1
1Developmental Neurobiology, Faculty of Biology and Biotechnology, Ruhr University Bochum, Bochum, Germany.
Abstract:
A battery of genetically encoded calcium indicators (GECIs) with different binding kinetics and calcium affinities was developed over the recent years to permit long-term calcium imaging. GECIs are calcium buffers and therefore, expression of GECIs may interfere with calcium homeostasis and signaling pathways important for neuronal differentiation and survival. Our objective was to investigate if the biolistically induced expression of five commonly used GECIs at two postnatal time points (days 14 and 22-25) could affect the morphological maturation of cortical neurons in organotypic slice cultures of rat visual cortex. Expression of GCaMP3 in both time windows, and of GCaMP5G and TN-XXL in the later time window impaired apical and /or basal dendrite growth of pyramidal neurons. With time, the proportion of GECI transfectants with nuclear filling increased, but an only prolonged expression of TN-XXL caused higher levels of neurodegeneration. In multipolar interneurons, only GCaMP3 evoked a transient growth delay during the early time window. GCaMP6m and GCaMP6m-XC were quite "neuron-friendly." Since growth-impaired neurons might not have the physiological responses typical of age-matched wildtype neurons the results obtained after prolonged developmental expression of certain GECIs might need to be interpreted with caution.
Insights
Certain genetically encoded calcium indicators (GECIs) can hinder neuronal development. Researchers found that GCaMP3, GCaMP5G, and TN-XXL impaired neuron growth, while others like GCaMP6m were neuron-friendly.
Area of Science:
- Neuroscience
- Cell Biology
- Biotechnology
Background:
- Genetically encoded calcium indicators (GECIs) are vital tools for long-term calcium imaging in neurons.
- As calcium buffers, GECIs may potentially disrupt neuronal calcium homeostasis, affecting differentiation and survival.
Purpose of the Study:
- To investigate the impact of biolistically expressed GECIs on the morphological maturation of cortical neurons.
- To assess effects at two distinct postnatal developmental time points (days 14 and 22-25) in rat visual cortex organotypic slice cultures.
Main Methods:
- Biolistic transfection of five common GECIs into rat visual cortex organotypic slice cultures.
- Analysis of neuronal morphology, including apical and basal dendrite growth, at postnatal days 14 and 22-25.
- Assessment of nuclear filling and neurodegeneration levels in GECI-expressing neurons.
Main Results:
- Expression of GCaMP3 (both time points), GCaMP5G, and TN-XXL (later time point) impaired pyramidal neuron dendrite growth.
- Prolonged TN-XXL expression led to increased neurodegeneration and nuclear filling.
- GCaMP6m and GCaMP6m-XC demonstrated minimal adverse effects on neuronal morphology, indicating they are 'neuron-friendly'.
Conclusions:
- Specific GECIs, including GCaMP3, GCaMP5G, and TN-XXL, can negatively affect cortical neuron maturation and survival.
- The choice of GECI and duration of expression are critical factors to consider during calcium imaging studies.
- Results highlight the need for caution when interpreting data from long-term calcium imaging experiments using certain GECIs.
