Abstract
Postsynaptic Ca2+ transients triggered by neurotransmission at excitatory synapses are a key signaling step for the induction of synaptic plasticity and are typically recorded in tissue slices using two-photon fluorescence imaging with Ca2+-sensitive dyes. The signals generated are small with very low peak signal/noise ratios (pSNRs) that make detailed analysis problematic. Here, we implement a wavelet-based de-noising algorithm (PURE-LET) to enhance signal/noise ratio for Ca2+ fluorescence transients evoked by single synaptic events under physiological conditions. Using simulated Ca2+ transients with defined noise levels, we analyzed the ability of the PURE-LET algorithm to retrieve the underlying signal. Fitting single Ca2+ transients with an exponential rise and decay model revealed a distortion of τrise but improved accuracy and reliability of τdecay and peak amplitude after PURE-LET de-noising compared to raw signals. The PURE-LET de-noising algorithm also provided a ∼30-dB gain in pSNR compared to ∼16-dB pSNR gain after an optimized binomial filter. The higher pSNR provided by PURE-LET de-noising increased discrimination accuracy between successes and failures of synaptic transmission as measured by the occurrence of synaptic Ca2+ transients by ∼20% relative to an optimized binomial filter. Furthermore, in comparison to binomial filter, no optimization of PURE-LET de-noising was required for reducing arbitrary bias. In conclusion, the de-noising of fluorescent Ca2+ transients using PURE-LET enhances detection and characterization of Ca2+ responses at central excitatory synapses.
Highlights
Single transmission events at excitatory synapses in the central nervous system elicit fast, short-lived rises in postsynaptic cytosolic [Ca2þ] (Ca2þ transients)
Ca2þ transients are readily detected with intracellular Ca2þ-sensitive dyes in minute (1–2 mm) structures such as dendritic spines, using two-photon laser-scanning fluorescence microscopy (TPLSM) [3,4,5]
TPLSM allows the optical recording of locally elicited excitatory postsynaptic Ca2þ transients (EPSCaTs) at single spines, simultaneously with the synaptic electrical response recorded at the soma [6]
Summary
Single transmission events at excitatory synapses in the central nervous system elicit fast, short-lived rises in postsynaptic cytosolic [Ca2þ] (Ca2þ transients). Ca2þ transients are readily detected with intracellular Ca2þ-sensitive dyes in minute (1–2 mm) structures such as dendritic spines, using two-photon laser-scanning fluorescence microscopy (TPLSM) [3,4,5] This technique offers excellent optical penetration and diffraction-limited excitation volume in structures embedded deep within brain tissue. TPLSM allows the optical recording of locally elicited excitatory postsynaptic Ca2þ transients (EPSCaTs) at single spines, simultaneously with the synaptic electrical response recorded at the soma [6] These techniques may be employed to determine how spine EPSCaTs encode specific patterns of synaptic activity that trigger or modulate synaptic plasticity [7,8,9,10,11]. The available information concerning EPSCaT time-course is limited, despite its importance [12,13]
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