Alveolar dynamics during respiration: are the pores of Kohn a pathway to recruitment?

Abstract

The change in alveolar size and number during the full breathing cycle in mammals remains unanswered, yet these descriptors are fundamental for understanding alveolar-based diseases and for improving ventilator management. Genetic and environmental mouse models are used increasingly to evaluate the evolution of disease in the peripheral lung; however, little is known regarding alveolar structure and function in the fresh, intact lung. Therefore, we have developed an optical confocal process to evaluate alveolar dynamics in the fresh intact mouse lung and as an initial experiment, have evaluated mouse alveolar dynamics during a single respiratory cycle immediately after passive lung deflation. We observe that alveoli become smaller and more numerous at the end of inspiration, and propose that this is direct evidence for alveolar recruitment in the mouse lung. The findings reported support a new hypothesis that requires recruitable secondary (daughter) alveoli to inflate via primary (mother) alveoli rather than from a conducting airway.

Figure 1.

Ex vivo mouse lung imaging schematic, consists of a custom iso-pressure system for inflating the lung to the desired pressure, a commercial Bio-Rad laser scanning confocal microscope, and a custom in vitro air- and watertight lung imaging chamber.

Figure 2.

Confocal images of the same mouse lung throughout an inflation/deflation cycle. Panels a through h represent 5-μm-thick laser scanning confocal microsopy sections from the same mouse lung inflated through pressures 0 to 35 cm H₂O in 5 cm H₂O increments, respectively; j through o show results for deflation. All images were acquired using a ×10 objective and field of view of 1.2 × 1.2 mm.

Figure 3.

Example of automated intercept labeling. Beginning of a wall is represented by a blue cross and end of a wall by a red cross. Logging of intercepts allows accurate calculation of airspace and wall chord lengths.

Figure 4.

(a) Change in lung volume versus pressure. (b) Alveolar airspace number in field of view (1.44 mm²) versus inflation pressure. (c) Mean chord length of alveolar airspace versus inflation pressure. (d) Mean chord length of alveolar walls. Error bars represent ± SD for five mice.

Figure 5.

(a) Inflation 0 to 35 cm H₂O, with wall intercepts. (b) Inflation 0 to 35 cm H₂O, clustered by color-coded area (μ²). (c) Inflation 0 to 35 cm H₂O, histogram of airspace chord lengths (μ). γ1 = Skew, γ2 = Kurtosis, red line = median value.

Figure 6.

(a) Deflation 35 to 0 cm H₂O, with wall intercepts. (b) Deflation 35 to 0 cm H₂O, clustered by color-coded area (μ²). (c) Deflation 35 to 0 cm H₂O, histogram of wall chord lengths (μ). γ1 = Skew, γ2 = Kurtosis, red line = median value.

Figure 7.

Two examples of alveoli “popping” open in C57BL/6 mouse lungs expressing green fluorescent protein. Each frame was captured over 50 milliseconds. Image was acquired using a catheter-based confocal microscopy technique.