Respiratory Physiology & Neurobiology 186 (2013) 197–205 Contents lists available at SciVerse ScienceDirect Respiratory Physiology & Neurobiology j our nal ho me p age: www.elsevier.com/locate/resphysiol Frontiers review Measurements of air ventilation in small vertebrates Jacopo P. Mortola a,∗ , Peter B. Frappell b a Department of Physiology, McGill University, Montreal, QC, Canada b University of Tasmania, Hobart, Tasmania 7001, Australia a r t i c l e i n f o Article history: Accepted 1 February 2013 Keywords: Airflow Barometric methodology Breathing pattern Control of breathing Infant monitoring Plethysmography Pneumotachography Respiratory techniques Tidal volume a b s t r a c t The breathing act is usually quite noticeable in most vertebrates; hence, the measurement of breathing frequency (f) rarely poses a serious problem. Differently, the quantitative assessment of tidal volume (V T ) (from which air ventilation, ˙ V E = f • V T , is computed) can be a major challenge. This article reviews the most common experimental approaches to quantify V T in adult or young vertebrates of small body size. In these animals, techniques commonly used in adult humans are unsuitable. Furthermore, physiologically meaningful data necessitate techniques with minimal disturbance to the subject under investigation. Dur- ing the last fifty years numerous and ingenious approaches have been developed and refined. Although none of them can be considered ideal or totally error-free, for specific tasks and/or species there is an optimal approach to measure tidal volume. © 2013 Elsevier B.V. All rights reserved. 1. Introduction The measurement of the quantity of air passing into the lungs over a period of time (minute ventilation, ˙ V E , the product of the air inhaled with each breath, tidal volume, V T , and of the num- ber of breaths per minute, breathing frequency, f) is part of the quantitative analysis of the breathing pattern, often necessary for clinical or research purposes. In air breathing vertebrates the f com- ponent of ˙ V E can be measured quite easily by visual inspection of the chest motion, as long as the respiration-related movements can be differentiated from locomotion and other non-respiratory movements of the chest. In many specimens, particularly those breathing fast or shallow like some small birds and mammals and other small-size vertebrates, visual counting of the breathing acts can be performed off-line during slow-speed play back of video recordings (e.g., Mortola and Limoges, 2006; Mortola and Seguin, 2009). Quite differently, the quantification of V T can pose serious challenges, especially in small animals. The measurement of V T represents the focus of this article. The simplest and most obvious approach to measure V T is to measure the volume of air exhaled into a leak-proof calibrated bag. This is what was done for many centuries, probably since almost two millennia ago, when the Greek philosopher and physician ∗ Corresponding author at: McGill University, McIntyre Medical Sciences Build- ing, Department of Physiology, Room 1121, 3655 Sir William Osler Promenade, Montreal, Quebec, Canada, H3G 1Y6. Fax: +1 514 398 7452. E-mail address: jacopo.mortola@mcgill.ca (J.P. Mortola). Galen collected expired air into a bladder. One likely source of error at the time must have been the change in temperature between subject and atmosphere. Gas laws were understood some fifteen centuries later and are now taken into consideration in the mea- surement of lung volumes. The spirometer, conceived in its original water-sealed version by John Hutchison in the middle of the nine- teen century (Kiraly, 2005), still represents the gold standard for the measurements of V T in adult humans, against which other techniques are compared for validation. However, in animals of small size the spirometer is impractical because of its substantial mechanical inertia; in addition, even in miniaturized format, the spirometer offers a significant dead space. Similarly, the recently introduced turbine wheels, the speed of which is proportional to airflow, are difficult to miniaturize and are known to underestimate the volume when flow rates are small. Over the years, several alter- native methodologies have been developed to quantify V T and ˙ V E in small vertebrates. The abundance of techniques testifies to the creativity and ingenuity of many investigators, but also indicates that none are universally accepted, ideal or error-free. This article presents some methodologies designed to quantify V T (and therefore ˙ V E ) in small animals, the size of a newborn human infant (3–4 kg) or less. Preference is given to familiar methodolo- gies most commonly used in conscious newborn or adult laboratory animals, noting that methods that involve anaesthesia or restraint (even visual disturbance) will affect the breathing pattern. This is not to imply that other approaches may not be better choices for particular experimental circumstances. Emphasis is placed on advantages and limitations; technical details can be traced from the representative references provided. 1569-9048/$ – see front matter © 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.resp.2013.02.001