Computer simulation of vocal tract resonance tuning strategies with respect to fundamental frequency and voice source spectral slope in singing.

A well-known concept of singing voice pedagogy is "formant tuning," where the lowest two vocal tract resonances ( fR1, fR2) are systematically tuned to harmonics of the laryngeal voice source to maximize the level of radiated sound. A comprehensive evaluation of this resonance tuning concept is still needed. Here, the effect of fR1, fR2 variation was systematically evaluated in silico across the entire fundamental frequency range of classical singing for three voice source characteristics with spectral slopes of -6, -12, and -18 dB/octave. Respective vocal tract transfer functions were generated with a previously introduced low-dimensional computational model, and resultant radiated sound levels were expressed in dB(A). Two distinct strategies for optimized sound output emerged for low vs high voices. At low pitches, spectral slope was the predominant factor for sound level increase, and resonance tuning only had a marginal effect. In contrast, resonance tuning strategies became more prevalent and voice source strength played an increasingly marginal role as fundamental frequency increased to the upper limits of the soprano range. This suggests that different voice classes (e.g., low male vs high female) likely have fundamentally different strategies for optimizing sound output, which has fundamental implications for pedagogical practice.

Are source-filter interactions detectable in classical singing during vowel glides?

In recent studies, it has been assumed that vocal tract formants (Fn) and the voice source could interact. However, there are only few studies analyzing this assumption in vivo. Here, the vowel transition /i/-/a/-/u/-/i/ of 12 professional classical singers (6 females, 6 males) when phonating on the pitch D4 [fundamental frequency (ƒo) ca. 294 Hz] were analyzed using transnasal high speed videoendoscopy (20.000 fps), electroglottography (EGG), and audio recordings. Fn data were calculated using a cepstral method. Source-filter interaction candidates (SFICs) were determined by (a) algorithmic detection of major intersections of Fn/nƒo and (b) perceptual assessment of the EGG signal. Although the open quotient showed some increase for the /i-a/ and /u-i/ transitions, there were no clear effects at the expected Fn/nƒo intersections. In contrast, ƒo adjustments and changes in the phonovibrogram occurred at perceptually derived SFICs, suggesting level-two interactions. In some cases, these were constituted by intersections between higher nƒo and Fn. The presented data partially corroborates that vowel transitions may result in level-two interactions also in professional singers. However, the lack of systematically detectable effects suggests either the absence of a strong interaction or existence of confounding factors, which may potentially counterbalance the level-two-interactions.

Quantifying syringeal dynamics in vitro using electroglottography.

The complex and elaborate vocalizations uttered by many of the 10,000 extant bird species are considered a major driver in their evolutionary success, warranting study of the underlying mechanisms of vocal production. Additionally, birdsong has developed into a highly productive model system for vocal imitation learning and motor control, where, in contrast to humans, we have experimental access to the entire neuromechanical control loop. In human voice production, complex laryngeal geometry, vocal fold tissue properties, airflow and laryngeal musculature all interact to ultimately control vocal fold kinematics. Quantifying vocal fold kinematics is thus critical to understanding neuromechanical control of voiced sound production, but in vivo imaging of vocal fold kinematics in birds is experimentally challenging. Here, we adapted and tested electroglottography (EGG) as a novel tool for examining vocal fold kinematics in the avian vocal organ, the syrinx. We furthermore imaged and quantified syringeal kinematics in the pigeon (Columba livia) syrinx with unprecedented detail. Our results show that EGG signals predict (1) the relative amount of contact between the avian equivalent of vocal folds and (2) essential parameters describing vibratory kinematics, such as fundamental frequency, and timing of syringeal opening and closing events. As such, EGG provides novel opportunities for measuring syringeal vibratory kinematic parameters in vivo Furthermore, the opportunity for imaging syringeal vibratory kinematics from multiple planar views (horizontal and coronal) simultaneously promotes birds as an excellent model system for studying kinematics and control of voiced sound production in general, including in humans and other mammals.

Japanese macaque phonatory physiology.

Although the call repertoire and its communicative function are relatively well explored in Japanese macaques (Macaca fuscata), little empirical data are available on the physics and the physiology of this species' vocal production mechanism. Here, a 6 year old female Japanese macaque was trained to phonate under an operant conditioning paradigm. The resulting 'coo' calls and spontaneously uttered 'growl' and 'chirp' calls were recorded with sound pressure level (SPL) calibrated microphones and electroglottography (EGG), a non-invasive method for assessing the dynamics of phonation. A total of 448 calls were recorded, complemented by ex vivo recordings on an excised Japanese macaque larynx. In this novel multidimensional investigative paradigm, in vivo and ex vivo data were matched via comparable EGG waveforms. Subsequent analysis suggests that the vocal range (range of fundamental frequency and SPL) of the macaque was comparable to that of a 7-10 year old human, with the exception of low intensity chirps, the production of which may be facilitated by the species' vocal membranes. In coo calls, redundant control of fundamental frequency in relation to SPL was also comparable to that in humans. EGG data revealed that growls, coos and chirps were produced by distinct laryngeal vibratory mechanisms. EGG further suggested changes in the degree of vocal fold adduction in vivo, resulting in spectral variation within the emitted coo calls, ranging from 'breathy' (including aerodynamic noise components) to 'non-breathy'. This is again analogous to humans, corroborating the notion that phonation in humans and non-human primates is based on universal physical and physiological principles.

A Set of Efficient nD NMR Protocols for Resonance Assignments of Intrinsically Disordered Proteins.

The RF pulse scheme RN[N-CA HEHAHA]NH, which provides a convenient approach to the acquisition of different multidimensional chemical shift correlation NMR spectra leading to backbone resonance assignments, including those of the proline residues of intrinsically disordered proteins (IDPs), is experimentally demonstrated. Depending on the type of correlation data required, the method involves the generation of in-phase ((15) N)(x) magnetisation via different magnetisation transfer pathways such as H→N→CO→N, HA→CA→CO→N, H→N→CA→N and H→CA→N, the subsequent application of (15) N-(13) C(α) heteronuclear Hartmann-Hahn mixing over a period of ≈100 ms, chemical-shift labelling of relevant nuclei before and after the heteronuclear mixing step and amide proton detection in the acquisition dimension. It makes use of the favourable relaxation properties of IDPs and the presence of (1) JCαN and (2) JCαN couplings to achieve efficient correlation of the backbone resonances of each amino acid residue "i" with the backbone amide resonances of residues "i-1" and "i+1". It can be implemented in a straightforward way through simple modifications of the RF pulse schemes commonly employed in protein NMR studies. The efficacy of the approach is demonstrated using a uniformly ((15) N,(13) C) labelled sample of α-synuclein. The different possibilities for obtaining the amino-acid-type information, simultaneously with the connectivity data between the backbone resonances of sequentially neighbouring residues, have also been outlined.

MAS solid state NMR of proteins: simultaneous ¹5N- ¹³CA and ¹5N- ¹³CO dipolar recoupling via low-power symmetry-based RF pulse schemes.

The generation of efficient RN n (ν)s,(ν)k symmetry-based low-power RF pulse schemes for simultaneous (15)N-(13)CA and (15)N-(13)CO dipolar recoupling is demonstrated. The method involves mixing schemes employing phase and amplitude-modulated dual band-selective 180° pulses as basic "R" element and tailoring of the RF field-modulation profile of the 180° pulses so as to obtain efficient magnetisation transfer characteristics over the resonance offset range of the nuclei involved. Mixing schemes leading to simultaneous (15)N-(13)CA and (15)N-(13)CO dipolar recoupling would permit the one-shot acquisition of different chemical shift correlation spectra that are typically utilized for protein backbone resonance assignments and thereby save data acquisition time. At representative MAS frequencies the efficacies of the mixing schemes presented here have been experimentally demonstrated via the simultaneous acquisition of {3D CONH and 3D CANH}, {3D CONH and 3D CO(CA)NH} and {3D CONH, 3D CANH, 3D CO(CA)NH and 3D CA(CO)NH} spectra generated via the magnetisation transfer pathways (1)H → (13)CO → (15)N → (1)H (CONH), (1)H → (13)CA → (15)N → (1)H (CANH) and (1)H → (13)CO → (13)CA → (15)N → (1)H (CO(CA)NH) and (1)H → (13)CA → (13)CO → (15)N → (1)H (CA(CO)NH).

HN-NCA heteronuclear TOCSY-NH experiment for (1)H(N) and (15)N sequential correlations in ((13)C, (15)N) labelled intrinsically disordered proteins.

A simple triple resonance NMR experiment that leads to the correlation of the backbone amide resonances of each amino acid residue 'i' with that of residues 'i-1' and 'i+1' in ((13)C, (15)N) labelled intrinsically disordered proteins (IDPs) is presented. The experimental scheme, {HN-NCA heteronuclear TOCSY-NH}, exploits the favourable relaxation properties of IDPs and the presence of (1) J CαN and (2) J CαN couplings to transfer the (15)N x magnetisation from amino acid residue 'i' to adjacent residues via the application of a band-selective (15)N-(13)C(α) heteronuclear cross-polarisation sequence of ~100 ms duration. Employing non-uniform sampling in the indirect dimensions, the efficacy of the approach has been demonstrated by the acquisition of 3D HNN chemical shift correlation spectra of α-synuclein. The experimental performance of the RF pulse sequence has been compared with that of the conventional INEPT-based HN(CA)NH pulse scheme. As the availability of data from both the HCCNH and HNN experiments will make it possible to use the information extracted from one experiment to simplify the analysis of the data of the other and lead to a robust approach for unambiguous backbone and side-chain resonance assignments, a time-saving strategy for the simultaneous collection of HCCNH and HNN data is also described.

Display glass for low-loss and high-density optical interconnects in electro-optical circuit boards with eight optical layers.

Parallel optical interconnects on-board level requires low propagation loss in wavelength range between 850 and 1550 nm to be compatible with datacom and telecom optical engines. For highest integration density tight waveguide bends and a scalable number of optical layers should be manufacturable for 2D interfaces to optical fiber array connectors and photonic assembly I/O's. We developed a glass waveguide panel process for double-sided processing of commercial available display glass by applying a two-step thermal ion-exchange process for low-loss multi-mode graded-index waveguides. Multiple glass waveguide panels can be embedded between electrical layers. The generic concept enables fabrication of high-density integration (HDI) electro-optical circuit boards (EOCB) with high number of optical and electrical layers. Waveguides with high NA of 0.3 for low bend losses could be achieved in glass with propagation loss of 0.05 dB/cm for all key wavelengths. Four of those glass waveguide panels were embedded in an EOCB demonstrator with size of 280 x 233 mm² providing eight optical layers with 96 channels in an area of 2.8 x 1.5 mm². To the best of our knowledge it's the highest number of layers that has ever been demonstrated for an EOCB.

An approach to NMR assignment of intrinsically disordered proteins.

An efficient approach to NMR assignments in intrinsically disordered proteins is presented, making use of the good dispersion of cross peaks observed in [(15) N,(13) C']- and [(13) C',(1) H(N) ]-correlation spectra. The method involves the simultaneous collection of {3D (H)NCO(CAN)H and 3D (HACA)CON(CA)HA} spectra for backbone assignments via sequential H(N) and H(α) correlations and {3D (H)NCO(CACS)HS and 3D (HS)CS(CA)CO(N)H} spectra for side-chain (1) H and (13) C assignments, employing sequential (1) H data acquisitions with direct detection of both the amide and aliphatic protons. The efficacy of the approach for obtaining resonance assignments with complete backbone and side-chain chemical shifts is demonstrated experimentally for the 61-residue [(13) C,(15) N]-labelled peptide of a voltage-gated potassium channel protein of the Kv1.4 channel subunit. The general applicability of the approach for the characterisation of moderately sized globular proteins is also demonstrated.

The softest sound levels of the human voice in normal subjects.

Accurate measurement of the softest sound levels of phonation presents technical and methodological challenges. This study aimed at (1) reliably obtaining normative data on sustained softest sound levels for the vowel [a:] at comfortable pitch; (2) comparing the results for different frequency and time weighting methods; and (3) refining the Union of European Phoniatricians' recommendation on allowed background noise levels for scientific and equipment manufacturers' purposes. Eighty healthy untrained participants (40 females, 40 males) were investigated in quiet rooms using a head-mounted microphone and a sound level meter at 30 cm distance. The one-second-equivalent sound levels were more stable and more representative for evaluating the softest sustained phonations than the fast-time-weighted levels. At 30 cm, these levels were in the range of 48-61 dB(C)/41-53 dB(A) for females and 49 - 64 dB(C)/35-53 dB(A) for males (5% to 95% quantile range). These ranges may serve as reference data in evaluating vocal normality. In order to reach a signal-to-noise ratio of at least 10 dB for more than 95% of the normal population, the background noise should be below 25 dB(A) and 38 dB(C), respectively, for the softest phonation measurements at 30 cm distance. For the A-weighting, this is 15 dB lower than the previously recommended value.

An approach to sequential NMR assignments of proteins: application to chemical shift restraint-based structure prediction.

A procedure for the simultaneous acquisition of {HNCOCANH & HCCCONH} chemical shift correlation spectra employing sequential [Formula: see text] data acquisition for moderately sized proteins is presented. The suitability of the approach for obtaining sequential resonance assignments, including complete [Formula: see text] and [Formula: see text] chemical shift information, is demonstrated experimentally for a [Formula: see text] and [Formula: see text] labelled sample of the C-terminal winged helix (WH) domain of the minichromosome maintenance (MCM) complex of Sulfolobus solfataricus. The chemical shift information obtained was used to calculate the global fold of this winged helix domain via CS-Rosetta. This demonstrates that our procedure provides a reliable and straight-forward protocol for a quick global fold determination of moderately-sized proteins.

Glottal opening and closing events investigated by electroglottography and super-high-speed video recordings.

Previous research has suggested that the peaks in the first derivative (dEGG) of the electroglottographic (EGG) signal are good approximate indicators of the events of glottal opening and closing. These findings were based on high-speed video (HSV) recordings with frame rates 10 times lower than the sampling frequencies of the corresponding EGG data. The present study attempts to corroborate these previous findings, utilizing super-HSV recordings. The HSV and EGG recordings (sampled at 27 and 44 kHz, respectively) of an excised canine larynx phonation were synchronized by an external TTL signal to within 0.037 ms. Data were analyzed by means of glottovibrograms, digital kymograms, the glottal area waveform and the vocal fold contact length (VFCL), a new parameter representing the time-varying degree of 'zippering' closure along the anterior-posterior (A-P) glottal axis. The temporal offsets between glottal events (depicted in the HSV recordings) and dEGG peaks in the opening and closing phase of glottal vibration ranged from 0.02 to 0.61 ms, amounting to 0.24-10.88% of the respective glottal cycle durations. All dEGG double peaks coincided with vibratory A-P phase differences. In two out of the three analyzed video sequences, peaks in the first derivative of the VFCL coincided with dEGG peaks, again co-occurring with A-P phase differences. The findings suggest that dEGG peaks do not always coincide with the events of glottal closure and initial opening. Vocal fold contacting and de-contacting do not occur at infinitesimally small instants of time, but extend over a certain interval, particularly under the influence of A-P phase differences.

Biomechanics of sound production in high-pitched classical singing.

Voice production of humans and most mammals is governed by the MyoElastic-AeroDynamic (MEAD) principle, where an air stream is modulated by self-sustained vocal fold oscillation to generate audible air pressure fluctuations. An alternative mechanism is found in ultrasonic vocalizations of rodents, which are established by an aeroacoustic (AA) phenomenon without vibration of laryngeal tissue. Previously, some authors argued that high-pitched human vocalization is also produced by the AA principle. Here, we investigate the so-called "whistle register" voice production in nine professional female operatic sopranos singing a scale from C6 (≈ 1047 Hz) to G6 (≈ 1568 Hz). Super-high-speed videolaryngoscopy revealed vocal fold collision in all participants, with closed quotients from 30 to 73%. Computational modeling showed that the biomechanical requirements to produce such high-pitched voice would be an increased contraction of the cricothyroid muscle, vocal fold strain of about 50%, and high subglottal pressure. Our data suggest that high-pitched operatic soprano singing uses the MEAD mechanism. Consequently, the commonly used term "whistle register" does not reflect the physical principle of a whistle with regard to voice generation in high pitched classical singing.

Complex vibratory patterns in an elephant larynx.

Elephants' low-frequency vocalizations are produced by flow-induced self-sustaining oscillations of laryngeal tissue. To date, little is known in detail about the vibratory phenomena in the elephant larynx. Here, we provide a first descriptive report of the complex oscillatory features found in the excised larynx of a 25 year old female African elephant (Loxodonta africana), the largest animal sound generator ever studied experimentally. Sound production was documented with high-speed video, acoustic measurements, air flow and sound pressure level recordings. The anatomy of the larynx was studied with computed tomography (CT) and dissections. Elephant CT vocal anatomy data were further compared with the anatomy of an adult human male. We observed numerous unusual phenomena, not typically reported in human vocal fold vibrations. Phase delays along both the inferior-superior and anterior-posterior (A-P) dimension were commonly observed, as well as transverse travelling wave patterns along the A-P dimension, previously not documented in the literature. Acoustic energy was mainly created during the instant of glottal opening. The vestibular folds, when adducted, participated in tissue vibration, effectively increasing the generated sound pressure level by 12 dB. The complexity of the observed phenomena is partly attributed to the distinct laryngeal anatomy of the elephant larynx, which is not simply a large-scale version of its human counterpart. Travelling waves may be facilitated by low fundamental frequencies and increased vocal fold tension. A travelling wave model is proposed, to account for three types of phenomena: A-P travelling waves, 'conventional' standing wave patterns, and irregular vocal fold vibration.

Phonovibrographic wavegrams: visualizing vocal fold kinematics.

Recently, endoscopic high-speed laryngoscopy has been established for commercial use as a state-of-the-art technique to examine vocal fold kinematics. Since modern cameras provide sampling rates of several thousand frames per second, a high volume of data has to be considered for visual and objective analysis. A method for visualizing endoscopic high speed videos in three-dimensional cycle-based graphs combining and extending the approaches of phonovibrograms and electroglottographic wavegrams is presented. To build a phonovibrographic wavegram, individual cycles of a phonovibrogram are segmented, normalized in cycle duration, and concatenated over time. For analyzing purposes, the emerging three-dimensional scalar field is visualized with different rendering techniques providing information of different aspects of vocal fold kinematics. The phonovibrographic wavegram incorporates information about the glottal closure type, size, and location of the amplitudes, symmetry, periodicity, and phase information. The potential of the approach to visualize the characteristics of vocal fold vibration in a compact and intuitive way is demonstrated within two healthy and three pathologic subjects. The phonovibrographic wavegram allows a comprehensive analysis of vocal fold kinematics and reveals information that remains hidden with other visualization techniques.

Voices in the ocean.

Toothed whales evolved a third way of making sounds similar to that of land mammals and birds.

Acoustical Theory of Vowel Modification Strategies in Belting.

Various authors have argued that belting is to be produced by "speech-like" sounds, with the first and second supraglottic vocal tract resonances (fR1 and fR2) at frequencies of the vowels determined by the lyrics to be sung. Acoustically, the hallmark of belting has been identified as a dominant second harmonic, possibly enhanced by first resonance tuning (fR1≈2fo). It is not clear how both these concepts - (a) phonating with "speech-like," unmodified vowels; and (b) producing a belting sound with a dominant second harmonic, typically enhanced by fR1 - can be upheld when singing across a singer's entire musical pitch range. For instance, anecdotal reports from pedagogues suggest that vowels with a low fR1, such as [i] or [u], might have to be modified considerably (by raising fR1) in order to phonate at higher pitches. These issues were systematically addressed in silico with respect to treble singing, using a linear source-filter voice production model. The dominant harmonic of the radiated spectrum was assessed in 12987 simulations, covering a parameter space of 37 fundamental frequencies (fo) across the musical pitch range from C3 to C6; 27 voice source spectral slope settings from -4 to -30 dB/octave; computed for 13 different IPA vowels. The results suggest that, for most unmodified vowels, the stereotypical belting sound characteristics with a dominant second harmonic can only be produced over a pitch range of about a musical fifth, centered at fo≈0.5fR1. In the [ɔ] and [ɑ] vowels, that range is extended to an octave, supported by a low second resonance. Data aggregation - considering the relative prevalence of vowels in American English - suggests that, historically, belting with fR1≈2fo was derived from speech, and that songs with an extended musical pitch range likely demand considerable vowel modification. We thus argue that - on acoustical grounds - the pedagogical commandment for belting with unmodified, "speech-like" vowels can not always be fulfilled.

Vocal Vibrato Characteristics in Historical and Contemporary Opera, Operetta, and Schlager.

OBJECTIVES/HYPOTHESIS: Vibrato is a core aesthetic element in singing. It varies considerably by both genre and era. Though studied extensively in Western classical singing over the years, there is a dearth of studies on vibrato in contemporary commercial music. In addressing this research gap, the objective of this study was to find and investigate common crossover song material from the opera, operetta, and Schlager singing styles from the historical early 20th to the contemporary 21st century epochs. STUDY DESIGN/METHODS: A total of 51 commercial recordings of two songs, "Es muss was Wunderbares sein" by Ralph Benatzky, and "Die ganze Welt ist himmelblau" by Robert Stolz, from "The White Horse Inn" ("Im weißen Rößl") were collected from opera, operetta, and Schlager singers. Each sample was annotated using Praat and analyzed in a custom Matlab- and Python-based algorithmic approach of singing voice separation and sine wave fitting novel to vibrato research. RESULTS: With respect to vibrato rate and extent, the three most notable findings were that (1) fo and vibrato were inherently connected; (2) Schlager, as a historical aesthetic category, has unique vibrato characteristics, with higher overall rate and lower overall extent; and (3) fo and vibrato extent varied over time based on the historical or contemporary recording year for each genre. CONCLUSIONS: Though these results should be interpreted with caution due to the limited sample size, conducting such acoustical analysis is relevant for voice pedagogy. This study sheds light on the complexity of vocal vibrato production physiology and acoustics while providing insight into various aesthetic choices when performing music of different genres and stylistic time periods. In the age of crossover singing training and commercially available recordings, this investigation reveals important distinctions regarding vocal vibrato across genres and eras that bear beneficial implications for singers and teachers of singing.

Dynamic System Coupling in Voice Production.

Voice is a major means of communication for humans, non-human mammals and many other vertebrates like birds and anurans. The physical and physiological principles of voice production are described by two theories: the MyoElastic-AeroDynamic (MEAD) theory and the Source-Filter Theory (SFT). While MEAD employs a multiphysics approach to understand the motor control and dynamics of self-sustained vibration of vocal folds or analogous tissues, SFT predominantly uses acoustics to understand spectral changes of the source via linear propagation through the vocal tract. Because the two theories focus on different aspects of voice production, they are often applied distinctly in specific areas of science and engineering. Here, we argue that the MEAD and the SFT are linked integral aspects of a holistic theory of voice production, describing a dynamically coupled system. The aim of this manuscript is to provide a comprehensive review of both the MEAD and the source-filter theory with its nonlinear extension, the latter of which suggests a number of conceptual similarities to sound production in brass instruments. We discuss the application of both theories to voice production of humans as well as of animals. An appraisal of voice production in the light of non-linear dynamics supports the notion that voice production can best be described with a systems view, considering coupled systems rather than isolated contributions of individual sub-systems.

Time-Synchronized MRI-Assessment of Respiratory Apparatus Subsystems-A Feasibility Study.

The thorax (TH), the thoracic diaphragm (TD), and the abdominal wall (AW) are three sub-systems of the respiratory apparatus whose displacement motion has been well studied with the use of magnetic resonance imaging (MRI). Another sub-system, which has however received less research attention with respect to breathing, is the pelvic floor (PF). In particular, there is no study that has investigated the displacement of all four sub-systems simultaneously. Addressing this issue, it was the purpose of this feasibility study to establish a data acquisition paradigm for time-synchronous quantitative analysis of dynamic MRI data from these four major contributors to respiration and phonation (TH, TD, AW, and PF). Three healthy females were asked to breathe in and out forcefully while being recorded in a 1.5-Tesla whole body MR-scanner. Spanning a sequence of 15.12 seconds, 40 MRI data frames were acquired. Each data frame contained two slices, simultaneously documenting the mid-sagittal (TH, TD, PF) and transversal (AW) planes. The displacement motion of the four anatomical structures of interest was documented using kymographic analysis, resulting in time-varying calibrated structure displacement data. After computing the fundamental frequency of the cyclical breathing motion, the phase offsets of the TH, PF, and AW with respect to the TD were computed. Data analysis revealed three fundamentally different displacement patterns. Total structure displacement was in the range of 0.94 cm (TH) to 4.27 cm (TD). Phase delays of up to 90∘ (i.e., a quarter of a breathing cycle) between different structures were found. Motion offsets in the range of -28.30∘ to 14.90∘ were computed for the PF with respect to the TD. The diversity of results in only three investigated participants suggests a variety of possible breathing strategies, warranting further research.