1 Answers
Fiber photometry is a calcium imaging technique that captures 'bulk' or population-level calcium activity from a specific cell-types within a brain region or functional network in order to study neural circuits This technique was originally designed to measure the afferent activity of projections to downstream brain regions in vivo. Population-level calcium activity can be easily correlated with behavioral tasks, such as spatial learning, memory recall and goal-directed behaviors.
Fiber photometry relies on the expression of genetically encoded calcium indicators , like GCaMP or RCaMP, which can be targeted to specific cells using cell-specific promoters like Ca2+/calmodulin-dependent protein kinase II and human synapsin that confer excitatory neuronal and pan-neuronal expression, respectively. These promoters can be used to target various neuronal subtypes as well as non-neuronal cells that exhibit calcium dynamics, such as astrocytes, using the glial fibrillary acidic protein promoter. In both neurons and astrocytes, cellular activity in the form of action potentials, exocytosis of neurotransmitters, changes in synaptic plasticity and gene transcription is coupled to an influx of Ca ions.
These activity-dependent changes in intracellular calcium levels can be monitored by introducing GECIs to the cell. Following this influx of ions, GECIs fluoresce upon Ca binding and the change in fluorescence corresponds proportionally to intracellular calcium changes. The most commonly used calcium indicator for fiber photometry is GCaMP6, although additional GECIs continue to be developed with unique fluorescence spectra, kinetics, signal-to-noise ratios and calcium-sensitivities. These indicators can be expressed in the brain in two main ways: viral expression and transgenic mouse lines. Recently, there has been a growing list of indicators that have become available to measure different chemical signals, like dLight to record dopamine signaling, or OxLight to record orexin, for example. GCaMP, RCaMP, dLight and other indicators are excited by a light source at an optimal wavelength and emit their own light in return, allowing for recording of calcium or neurotransmitter dynamics across time.
Fiber photometry systems are designed to deliver precise excitation wavelengths of light that are specific to a calcium or neurotransmitter indicator. This light travels down an optical fiber to a fiber optic that is implanted in the brain region or regions of interest. The calcium indicator is that is expressed in a cell-type specific manner is excited by this light and in turn, emits its own signal that travels back through the same fiber. These collected emission signals are spectrally-separated by a dichroic mirror, passed through a filter and focused onto a photodetector, scientific camera, or PMT. The collected signal represents a change in fluorescence relative to an initial baseline. In turn, researchers can observe a signal that corresponds to calcium transients. This time series data can be analyzed using a variety of open-source pipelines, such as pMAT, pyPhotometry and GuPPy.