Muscle architecture of the forelimb of the Golden Pheasant (Chrysolophus pictus) (Aves: Phasianidae) and its implications for functional capacity in flight

Abstract

Background

Flight is the central avian adaptation in evolution. Wing muscles form an important anatomical basis for avian flight, affecting wing performance and determine modes of flight. However, the roles of distal muscles in adjusting the wing, as well as their functional specializations, remain largely unknown. The importance of muscle fiber architecture has long been recognized. In this study, we provide quantitative anatomical data on the muscle architecture of the forelimb of the Golden Pheasant (Chrysolophus pictus), with an emphasis on brachial, antebrachial and manual segments.

Methods

The forelimbs of five Golden Pheasants were dissected and detailed measurements of all muscles were made, including muscle mass, muscle belly length, fascicle length. From these values, muscle volume, physiological cross-sectional area (PCSA) and maximum isometric force were derived.

Results

General trends such as the distribution of muscle mass, fascicle length and the ratio of tendon length/belly length are revealed. Comparing PCSAs between antebrachial depressors and elevators and between intrinsics of the alular digit and major digit yielded significant differences (p < 0.05). Pronounced development of the antebrachial depressors suggests that ventral rotation of the distal half of the wing is a pivotal factor in shape change and orientation modulation. Large PCSAs in tandem with the force generation capability of the major digit intrinsics may help stabilize the digits while enhancing support of the primary feathers. The architectural properties of the alular digit confirm that alular adjustment is essential to rapid adduction and abduction.

Conclusions

These observations illustrate the underlying structural basis for the functional capacities of the distal forelimb muscles and may provide additional information useful in further biomechanical and in vivo investigations.

Background

Flight is the central avian adaptation and an extraordinary range of specialized modes of flight has been exploited during evolution (Gill 1995). Skeleton and wing muscles form the basis for avian flight; these anatomical features are also adapted to reflect the flight style and phylogeny (McKitrick 1991; Corvidae et al. 2006). Muscle architecture and the physiological properties of muscle fiber are important in the evaluation of muscle capacity. With few exceptions, muscle mass and volume distribution are considered key physical parameters in avian myology, controlling flight (Fisher 1946; Owre 1967). Additional studies are needed to investigate the relationship between fiber-type distribution and muscle function (e.g., Meyers 1992; Torrella et al. 1998; Meyers and Stakebake 2005; Corvidae et al. 2006; Welch and Altshuler 2009). Mass, volume and the type of fiber substantially affect the contractile properties of a muscle. However, more than any other factor, muscle architecture predicts muscle function (Lieber and Fridén 2000; Ward et al. 2009; Lieber and Ward 2011). The importance of muscle fiber architecture has long been recognized and related studies have been widely reported in human and other mammals (Sacks and Roy 1982; Payne et al. 2005, 2006; Williams et al. 2007, 2008; Eng et al. 2008; Channon et al. 2009; Rupert et al. 2014), but seldom in birds. Report on the hind limb of an ostrich by Smith et al. (2006) may have been the first comprehensive research on avian muscle architecture. More work, with a special focus on the functional relationship between muscle architecture and flight styles, is needed (Dial 1992a).

Skeletal muscle architecture is defined as the arrangement of muscle fibers within a muscle (Gans 1982; Sacks and Roy 1982). The two most important architectural parameters are the physiological cross-sectional area (PCSA) and muscle fiber length (Lieber and Fridén 2000). The PCSA of a muscle is the only architectural parameter that is directly proportional to force generation, whereas muscle excursion and velocity are directly proportional to muscle fiber length. A greater serial sarcomere count (i.e., greater fiber length) leads directly to a larger muscle excursion, because serial excursions of individual sarcomeres are additive (Lieber and Ward 2011). Thus, long fibers are predicted to operate over a relatively wide range of muscle lengths, which can achieve greater velocities compared with shorter-fibered muscles. Due to these direct structure-function correlations, architectural features will undoubtedly reflect the functional properties and specializations of different muscles.

The pectoralis and supracoracoideus play dominant roles in bird flight and are the most widely studied of the forelimb muscles. Electromyographic (EMG) data indicate that intrinsic muscles of the wing contribute little additional mechanical power for flight, but are important in modulating wing orientation and controlling wing shape (Dial 1992a, 1992b; Biewener 2011). Due to the difficulty of in vivo force measurements for smaller muscles located more distally in the wing, the roles of these muscles in adjusting the wing, as well as their functional specializations, remain largely unknown (Biewener 2011). Given this circumstance, analysis of muscle architecture can play an important role in evaluating the role of muscles. China is particularly rich in pheasants and their relatives, with the result that these receive more attention than any other group. The Golden Pheasant (Chrysolophus pictus) is an endemic pheasant of China, but with their number decreasing (Lei and Lu 2006). The flight of this species, as well as that of other phasianids involves an explosive take-off followed by a rapid and swift flapping flight (Askew and Marsh 2002). Many investigations have focused on the power output and function of wing muscles during non-steady flight (Dial 1992a, 1992b; Tobalske and Dial 2000; Askew et al. 2001; Askew and Marsh 2002). In this study we have attempted to quantify forelimb muscles architecture in the Golden Pheasant, with an emphasis on the brachial, antebrachial and manual segments and their varying functions in adjusting wing shape. An understanding of muscle architectural specialization illuminates the functional features of the different muscles used during flight and may provide additional information for further biomechanical and in vivo investigations.

Methods

Five adult Golden Pheasants [three females, two males; mass 422 ± 95.8 g (mean ± SD)] were obtained from an accredited local farm for use in this study, killed by decapitation. The project was approved by the Animal Care and Ethics Committee of Capital Normal University. After the birds were sacrificed, each limb was skinned and individual muscles were identified, exposed and cleared of fascia. The muscles were then systematically removed and detailed dissections of muscle-tendon architecture were carried out. Muscle architecture was determined from methods described by Smith et al. (2006). Muscle mass measurements were obtained with an electronic scale (Ohaus, USA) to the nearest 0.0001 g, while lengths were measured with digital calipers (Workzone, Germany) to the nearest 0.01 mm. Muscle belly length was measured as the length from the most proximal fibers to the most distal fibers. Muscle volume was calculated by dividing muscle mass by muscle density (1.06 g∙cm⁻³; Mendez and Keys 1960; Brown et al. 2003) and its physiological cross-sectional area (PCSA) was then determined by dividing muscle volume by its mean fascicle length. To obtain fascicle length, the collagen between the muscle fibers was gradually dissolved in nitric acid (30% HNO₃) for about 24 h and then the tissue was immersed in a 50% glycerol solution. At least five measurements of fascicle length were taken from randomly distributed areas and depths within the muscle belly. The maximum isometric force of a muscle, F _max, was estimated by multiplying PCSA by the maximum isometric stress of a vertebrate skeletal muscle (0.3 MPa; Wells 1965). Pennation angles were not included in our measurements, because muscle bellies in most of the forelimb muscles are directly attached to the skeleton, without forming tendons of insertion. As well, among muscles with an insertion tendon, nearly all angles are smaller than 10°; the cosine of a 10° angle is very close to one and would thus have little effect on estimations of PCSA.

For gross comparison, muscles were assigned to six functional groups: i.e., brachial depressors/elevators, antebrachial extensors/flexors, antebrachial depressors/elevators, manual extensors/flexors, manual depressors/elevators and manual intrinsics. For each functional muscle group, the sum of the separate muscle PCSAs and the total force were combined to result in one value for each group; the fascicle length was averaged among the containing muscles of that group. One way ANOVA was performed within each functional group. Functions of individual muscles have not been previously published for this species and are, therefore, based on anatomical positioning, our personal observations and references from George and Berger (1966).

Results

General features

All 47 previously recognized muscles (Zhang and Yang 2013) were identified in the pectoral limb of the Golden Pheasant. The corresponding architectural data are presented in Table 1. On average, the unilateral forelimb muscle mass of the Golden Pheasant accounts for 12.00% ± 1.56% (mean ± SD) of body mass, corresponding to 51.74 g of muscle per forelimb. Muscles were assigned to four groups according to their location; the distribution of muscle mass exhibited a sharp reduction from the proximal to the distal (Figure 1). Extrinsic muscles, brachial muscles, antebrachial muscles and manual muscles constitute 84.98%, 8.81%, 5.94% and 0.33%, respectively, of the total forelimb muscle mass. The pectoralis (PT), supracoracoideus (SP) and scapulohumeralis caudalis (SHC) are the three largest muscles of the forelimb. PT accounts for 13.4% of the total body mass and 55.86% of the total forelimb muscle mass. The corresponding data for SP are 3.9% and 16.31% and for SHC 1.3% and 5.33%. The largest brachial muscle was triceps brachii, accounting for 4.63% of the total forelimb muscle mass, closely followed by biceps brachii (BB, 1.66%) and deltoideus major (DMA, 1.41%). Triceps brachii is composed of two distinct heads, the humeral and scapular, with the humeral head (TH, 1.331 ± 0.490 g) slightly larger than the scapular head (TS, 1.076 ± 0.439 g). The extensor metacarpi radialis (EMR) and flexor carpi ulnaris (FCU) muscles form the bulk of the antebrachial group, weighing 0.614 ± 0.284 g and 0.469 ± 0.293 g, respectively, corresponding to 1.17 and 0.87% of the total forelimb muscle mass. Smallest were the distal manual muscles, which include the flexor alulae (FA, 0.004 ± 0.002 g, 0.008%), the extensor brevis alulae (EBA, 0.007 ± 0.005 g, 0.014%) and the flexor digiti minoris (FDMI, 0.011 ± 0.006 g, 0.020%).

Table 1 Architectural properties of forelimb muscles in the Golden Pheasant

Figure 1

Mean muscle mass (± SD) as a proportion of total forelimb muscle mass for all muscles. The pectoralis is omitted here because of its large percentage (55.86%) of the forelimb of the Golden Pheasant.

Among muscles with an insertion tendon, tendon lengths were obtained only from those that were discernible. The proportional lengths of muscle belly and tendon lengths are shown in Figure 2. Most tendons in the distal limb are comparatively longer than those in the proximal limb and exceed belly lengths.

Figure 2

Comparison between tendon length and muscle belly length for both the proximal (dark red and dark blue) and distal (light red and light blue) forelimb muscles of the Golden Pheasant.

The distribution of fascicle lengths showed a general trend, with proximal muscles having longer fascicles and distal muscles relatively short fascicles. The pectoralis and supracoracoideus had the longest mean fascicle lengths (2.44 cm and 2.33 cm, respectively), followed closely by the latissimus dorsi, deltoideus major, tensor propatagialis and scapulohumeralis caudalis. Intrinsic manual muscles displayed the shortest fascicle lengths, ranging from 0.16 to 0.44 cm. Among the intrinsics, the abductor and adductor of the alular digit were relatively high in this parameter.

The pectoralis muscle had, on average, the largest PCSA (11.15 cm²), thus yielding the highest force-producing capacity of all the forelimb muscles (F _max = 334.55 N), followed by the supracoracoideus (3.63 cm², 108.95 N). PCSAs of most of the intrinsics of the manus, i.e., latissimus dorsi pars cranialis, scapulohumeralis cranialis, serratus superficialis pars cranialis and brachialis, were the smallest, at less than 0.1 cm².

Comparisons between functional groups

The fascicle length and PCSA of six functional groups are shown in Table 2. A significant difference (p < 0.05) was observed between the antebrachial depressors and elevators for PCSA and total force. The difference in PCSA and total force between the intrinsics of the alular digit and major digit was also significant (p < 0.05).

Table 2 Comparisons of muscle group architecture

Disparities in architectural properties and mechanical function within a synergic group, or between different functional groups, are shown in Figure 3. The antebrachial extensors were characterized by high force-producing capacities, whereas the flexors showed more diversification in their fundamental design: FCU and BB would be expected to govern function because of their large PCSAs, while TP has the capacity to perform a large excursion. Considering the depressors (or pronators) vs. elevators (supinators) comparison, the other antagonistic groups of the antebrachium, the former possessed a significantly larger PCSA and relatively longer FL than the latter. The FCU appeared to control the flexion of the manus in force production. Extension of the manus tended to be more complicated than flexion for simultaneously possessing a large PCSA muscle (EMR) and a long FL muscle (ELA), features that suggest a design predicated on both force and speed. All intrinsic muscles of the manus are indicated on the bottom of the left-hand side of Figure 3 (low PCSA, short fascicles). They differ anatomically in that the muscles of the major digit are characterized by large PCSAs, whereas the muscles in the alular digit are remarkable for their long fascicle lengths.

Figure 3

Scatter graph of fascicle length and physiological cross-sectional areas (PCSAs) of muscles in the forelimb of the Golden Pheasant.

Discussion and conclusions

Along with certain mammals and birds, such as horses and ostriches (Payne et al. 2005; Smith et al. 2006), the distribution of muscle mass throughout the Golden Pheasant forelimb demonstrated a proximal-to-distal reduction. This design minimizes the moment of inertia during locomotion (Hildebrand 1988), which, in turn, conserves metabolic energy (Steudel 1996). The tendon in the distal forelimb segment of the Golden Pheasant was relatively longer than that found in the proximal. This extra length enables muscles to control distal movements of the wing without the burden of extra muscle mass. Possessing a small and lightweight distal segment (Biewener 2011) provides a distinct advantage by decreasing limb inertia during flight. The ability of tendons to stretch and recoil enables storage and recovery of elastic energy, while allowing muscle fibers to sustain high forces (Roberts 2002). Longer tendons in the distal forelimb enhance muscle performance by increasing contraction efficiency and reducing metabolic costs.

As the primary wing depressor and elevator, the pectoralis and supracoracoideus are the most widely studied of the forelimb muscles. In the Golden Pheasant, these two muscles possess the largest mass, PCSA and proportion of all the muscles in the forelimb. As expected, they also possess the longest fascicle lengths. Greenwalt (1962) predicted that the pectoralis muscle of a volant species should constitute 15.5% of the total body mass of a bird. In the Golden Pheasant, this muscle comprises 13.4% of total body mass. Regarding the supracoracoideus, this muscle represents about 1.6% of the total body mass in volant birds and most non-diving birds, 4%–5% in wing-propelled diving birds (e.g., Atlantic Puffin) and 10%–12% in penguins (Greenwalt 1962; Poore et al. 1997; Kovacs and Meyers 2000). Our results show that the supracoracoideus in the Golden Pheasant accounts for 3.9% of the total body mass of this species. The relatively large supracoracoideus in alcids and other wing-propelled diving birds most likely evolved to raise the wing against the resistive drag of water (Kovacs and Meyers 2000). The pheasants use high-frequency, high-amplitude wing beats during their explosive take-off flights and these attributes probably create a high inertial power requirement for elevating the wing. In the Golden Pheasant, it is likely that the supracoracoideus is large to meet this inertial-work or inertial-power requirement. A long fascicle results in greater excursion length, whereas PCSA corresponds to force production. The long fascicle and large PCSA of the pectoralis and supracoracoideus enable powerful upstrokes and downstrokes through a large excursion to achieve sufficient aerodynamic lift, particularly during takeoff and vertical ascending flights (Biewener 2011).

Previous work (Dial 1992a; Berg and Biewener 2010) demonstrated that the brachial and antebrachial muscles act primarily as joint stabilizers and are not essential for normal extension and flexion of the wing during level flapping flights. However, during non-steady flights (e.g., takeoffs and landings), these muscles contribute to the performance of the wing by acting as an aerofoil, modulating wing orientation and wing shape (Dial 1992a, 1992b; Biewener 2011). In this study, we have demonstrated differences in architectural design within antagonistic groups of the antebrachial muscles (Table 2). For example, antebrachial elevators vs. depressors present significant differences in total PCSA and estimated isometric force (p < 0.05). Similar differences were also observed between intrinsic muscles of the major and alular digits, which have architectural features that facilitate force production (major digits) and excursion (alular digit). Pronounced development of the antebrachial depressors (pronator superficialis, pronator profundus and ectepicondylo ulnaris) suggests that ventrally rotating the distal half of the wing should profoundly affect shape change and orientation modulation during non-steady flights; these muscles are evidently capable of providing increased thrust for acceleration during takeoff and vertical ascending flights (Biewener 2011). A similar trend was also found in pigeons, where the pronator superficialis exhibited biphasic activities with EMG intensities at their maximum during takeoff and ascending flights (Dial 1992b). Three birds of prey (Cooper’s Hawk Accipiter cooperii, Osprey Pandion haliaetus and Red-tailed Hawk Buteo jamaicensis) also exhibit these muscle-induced phenomena, which may explain observed differences in flight mode and hunting behavior (Corvidae et al. 2006). The manus intrinsics (Figure 3) may be related to joint stabilization or the execution of precision movements (Williams et al. 2008; Channon et al. 2009). The major digit provides support for the outer primaries that control forward thrust and aerodynamic performance, particularly during flapping flights (Combes and Daniel 2001; Swaddle and Lockwood 2003). The major digit intrinsics, with their greater PCSAs and force generation capability, may help stabilize the wing and provide powerful support for the primary feathers. The alula, or bastard wing, is a high lift device located at the leading edge of the wing that allows birds to fly at an acute attack angle and at a lower speed without stalling (Gill 1995; Alvarez et al. 2001). It features 3 to 5 small flight feathers originating from the first digit and moves independently of the rest of the wingtip. Four muscles attach to the alular digit and control the position of the alula; among them, the adductor and abductor appear to be dominant and more effective functionally due to their significantly longer fascicles. Their architectural properties are designed for excursion and velocity, as muscles with relatively long fibers operate over a large range of muscle lengths and can achieve faster velocities compared with a shorter-fibered muscle (Ward et al. 2009). This may imply that adjustment of the alula is paramount for rapid adduction and abduction during flight in the Golden Pheasant.

Our results also revealed that muscle architecture varies widely within synergic groups (Figure 3, Table 2). The flexors of the manus, for example, are composed of six different muscles: i.e., flexor carpi ulnaris, flexor digitorum profundus, flexor digitorum superficialis, extensor carpi ulnaris, ulnometacarpalis ventralis and ulnometacarpalis dorsalis. They all contribute to flexion of the manus. The flexor carpi ulnaris (FCU), with the largest PCSA at nearly 8 times that of the flexor digitorum superficialis, generates very high forces and acts as the functionally dominant muscle in this group. Within the manual extensors group, extensor metacarpi radialis was designed for optimal force production, whereas extensor longus alulae evolved for fast velocity. These results are consistent with Dial’s observation (1992a) that the extensor metacarpi radialis and flexor carpi ulnaris exhibited their greatest EMG activity during non-steady flights, indicating that the manus extension is significantly more complicated than flexion.