Back to article: Exocytotic fusion pore under stress
FIGURE 3: Cell-attached membrane capacitance (Cm) measurements on pituitary lactotrophs show transient (reversible) and full fusion exocytotic events.(A) Lactotrophs co-transfected with hyperpolarization-activated and cyclic nucleotide-gated 2 pDNA and EGFP pDNA were used for the Cm measurements in the cell-attached configuration [12]. (Ai–Aiii) All measurements were performed only on lactotrophs exhibiting EGFP fluorescence, as shown for the same cell under transmitted light (Ai), under epifluorescence (Aii), and during the measurement (Aiii). Scale bar, 3 μm. (B) An epoch from a representative recording showing the reversible steps in the Im part of the admittance trace. Reversible events in Im, which likely represent transient exocytotic events (transient fusion pore opening), either exhibited projections to the Re part of the admittance trace (1) or not (2). The arrow points to the calibration pulse in Im, which was triggered manually to ensure the correct phase angle settings. (C) Calculated vesicle capacitance (Cv) and fusion pore conductance (Gp), a measure of fusion pore geometry, for all reversible events with projections [74]. (D) A representative example of irreversible upward steps in the Im trace and the corresponding Re trace (3), which likely denotes a full fusion exocytotic event. (E) A scheme of the stages that a vesicle must pass to completely fuse with the plasma membrane: from narrow fusion pore formation (1), to widening of the fusion pore (2), and finally to full fusion of the vesicle membrane and the plasma membrane (3). From Ref. [74].
12. Rituper B, Gucek A, Jorgacevski J, Flasker A, Kreft M, Zorec R (2013). High-resolution membrane capacitance measurements for the study of exocytosis and endocytosis. Nat Protoc 8(6): 1169-1183. 10.1038/nprot.2013.069
74. Calejo AI, Jorgacevski J, Rituper B, Gucek A, Pereira PM, Santos MA, Potokar M, Vardjan N, Kreft M, Goncalves PP, Zorec R (2014). Hyperpolarization-activated cyclic nucleotide-gated channels and cAMP-dependent modulation of exocytosis in cultured rat lactotrophs. J Neurosci 34(47): 15638-15647. 10.1523/JNEUROSCI.5290-13.2014