Optimal Pipette Resistance, Seal Resistance, and Zero-Current Membrane Potential for Loose Patch or Breakthrough Whole-Cell Recording in vivo

In vivo loose patch and breakthrough whole-cell recordings are useful tools for investigating the intrinsic and synaptic properties of neurons. However, the correlation among pipette resistance, seal condition, and recording time is not thoroughly clear. Presently, we investigated the recording time of different pipette resistances and seal conditions in loose patch and breakthrough whole-cell recordings. The recording time did not change with pipette resistance for loose patch recording (R(p)-loose) and first increased and then decreased as seal resistance for loose patch recording (R(s)-loose) increased. For a high probability of a recording time ≥30 min, the low and high cutoff values of R(s)-loose were 21.5 and 36 MΩ, respectively. For neurons with R(s)-loose values of 21.5–36 MΩ, the action potential (AP) amplitudes changed slightly 30 min after the seal. The recording time increased as seal resistance for whole-cell recording (R(s)-tight) increased and the zero-current membrane potential for breakthrough whole-cell recording (MP(zero-current)) decreased. For a high probability of a recording time ≥30 min, the cutoff values of R(s)-tight and MP(zero-current) were 2.35 GΩ and −53.5 mV, respectively. The area under the curve (AUC) of the MP(zero-current) receiver operating characteristic (ROC) curve was larger than that of the R(s)-tight ROC curve. For neurons with MP(zero-current) values ≤ −53.5 mV, the inhibitory or excitatory postsynaptic current amplitudes did not show significant changes 30 min after the seal. In neurons with R(s)-tight values ≥2.35 GΩ, the recording time gradually increased and then decreased as the pipette resistance for whole-cell recording (R(p)-tight) increased. For the high probability of a recording time ≥30 min, the low and high cutoff values of R(p)-tight were 6.15 and 6.45 MΩ, respectively. Together, we concluded that the optimal R(s)-loose range is 21.5–36 MΩ, the optimal R(p)-tight range is 6.15–6.45 MΩ, and the optimal R(s)-tight and MP(zero-current) values are ≥2.35 GΩ and ≤ −53.5 mV, respectively. Compared with R(s)-tight, the MP(zero-current) value can more accurately discriminate recording times ≥30 min and <30 min.