The sonic muscle of toadfish is the fastest vertebrate muscle ever measured, and the rate of transport of Ca2+ and dissociation of cross-bridge are also fatest. Parvalbumins are Ca2+-binding proteins present in vertebrate muscle, and they can aid muscle relaxation. Several isoforms of parvalbumin had been identified and presented in variable proportion in different kinds of muscles (e.g. red muscle, white muscle and pink muscle). Both male and female grunting toadfish (Allenbatrachus grunniens) have intrinsic sonic muscles attached on swim bladders. The morphology of male and female sonic muscle was compared, and no significant differences in both length, width, thickness and weight were found. SDS-PAGE and western blotting were used to determine the total parvalbumin expression and identify the parvalbumins from sonic muscle and body white muscle. There were no significant differences in total parvalbumin expression in sonic muscle and body white muscle. The result indicates that there is no positive correlation between high content of parvalbumins and speed of muscle relaxation. In native-PAGE, two and four parvalbumin isoforms were identified from sonic muscle and body white muscle, respectively. The estimated size of Parv1, Parv2 and Parv3 size in grunting toadfish¡¦s sonic muscle were 10kDa, 10.5kDa and 10.5kDa, respectively, and the isoelectric points of Parv1, Parv2 and Parv3 in grunting toadfish were 4.77, 4.58 and 4.42, respectively. In the sonic muscle, the major parvalbumin isoform was parvalbumin isoform 1 (Parv1), which comprised more than 94% of total parvalbumin, and parvalbumin isoform 2 (Pav2) comprised vi only 5% of total parvalbumin content. In body white muscle, on the other hand, the major isoform was parvalbumin isoform 2 (Parv2) which comprised 58% of toal parvalbumin. Both Parv1 (with Parv1a) and Parv3 comprised about 20%. Parvalbumin isoforms were be discussed. The result supports that Parv1 has a highest effect on the relaxation of the grunting-toadfish¡¦s sonic muscle.

The swim bladder of Glaucosoma hebracium is specialized by having two side membranes extending dorsally from the side to the vertebrae. This characteristic supports the morphology of the Glaucosomatidae. There is a pair of reddish muscles connecting the skull and the swim bladder of the Glaucosoma hebracium. This type of muscle is likely to involve with sound production. However, vocalization has not been reported in the Glaucosomatids. As direct recordings for sound production in live specimens of this species could not easily be carried out, function of these swim bladder muscles was assessed by means of biochemical assay. Activities of Citrate synthase (CS) and L-lactate dehydrogenase (LDH), known to involve with energy supply during muscle contraction were assessed. Two-dimensional gels for the white muscle, tail region red muscle and swim bladder muscles were compared to reveal difference of protein expression. Results show that CS activity of the swim bladder muscle was higher than the white muscle. But LDH activities in these two types of muscles were not significant different. As high activities in CS reflecting aerobic nature of the tissue, the present dataset suggests that the swim bladder muscles should involve with sound production. Proteomics analyzes identified 14 proteins from the swim bladder muscle, one of them are related to energy production (ATP synthase beta chain). Two-dimensional gels show some different spots from these three type muscles. These results help us to find some different proteins in these three types of muscles of this fish. In conclusions base on these evidence in morphology, histology, enzyme activity assay and two-dimensional gels, the swim bladder muscles should involve with sound production in G. hebracium.

Batrachoididae, one of the most well-studied soniferous fishes, are typical examples of fish using intrinsic sonic muscles to excite vibration of swim bladder to emit sound. Most fishes possess a single swim bladder including the Batrachoididae. However, the grunting toadfish (Allenbatrachus grunniens) was found to have two separated swim bladders located in the dorsal part of the abdominal cavity; the size of the swim bladders were quite similar. Sonic muscles were firmly attached to the lateral side of each swim bladder. The aims of this study were to (1) investigate the specialized swimbladder morphology and the acoustic signals of the grunting toadfish (A. grunniens), (2) to falsify the hypothesis that this unique form of swim bladder is a synapomorphic character, and species that possess this character are sister groups. The vocal repertoire in grunting toadfish can generally divided into two types ¡V grunt and boatwhistle. Grunts were harmonic signals with shorter call duration, and could be emitted alone as single grunt (i.e., hand-held grunt) or in series (known as grunt train). Boatwhistles were also hamonic but much longer in call duration and usually appeared in succession. A small portion of signals were found to have acoustic beats, which was previously described in the three-spined toadfish (Batrachomoeus trispinosus), which also possess a pair of swim bladders. Therefore, signals with acoustic beats may be a key character for generating sound by two separated swim bladders. Comparing the morphological measurements of swim bladders indicated that there were no significant differences between swim bladders on different sides. However, comparisons between genders showed that the width, thickness, and weight of sonic muscle in females were significantly higher than males. Less wide sonic muscles with shorter sonic muscle fibers may enables the muscle to contract at a higher velocity in male fishes. However, females were found to have thicker sonic muscle, which indicated that the vocal ability in females may be higher than it was expected. The sonic muscle fibers of females have a larger myofibrillar region compared with males, which have a relatively larger central core. These characters may increase the fatigue-resistance of sonic muscle in males, which can contract at a relatively higher rate for a longer duration. Species with two separated swim bladders were found to be sister group in the molecular phylogenetic tree, implying that this specialized morphological character is synapomorphy.

Abstract Sciaenids are a kind of economic oceanic fish with the most dynamic vocal activity. It is important to find out what type of sound a soniferous fish can make. Therefore, I used this acoustic character as a tool to study the distribution of sciaenids in order to manage their fisheries. In the previous researches, scientist found that there were eight types of sounds (A~H type) appearing on the coastal areas of Yunlin, and the frequency range of the H-type sound could reach to 8000Hz. This sound was suspected to be made by Otolithes ruber. Connaughton (1994) and Sprague (2000) mentioned that the differences of length, weight, and tension of the sonic muscle and the amount of protein and glycogen affect the types of sounds emitted. To reveal the producer of high-frequency sound, this research analyzed and compared the length, width, thickness and somatic of sonic muscle and the amount of protein and glycogen in seven sciaenid species occuring on the west coast of Taiwan (Johnius tingi, Johnius sina, Johnius amblycephalus, Johnius amblycephalus, Pennahia argentata, Chrysochir aureus, and Otolithes ruber), then to find if the differences of physic parameter and energy supplication are the major factors making O. ruber produce high frequency of sound. In this research, I found that the length of O. ruber¡¦s sonic muscle was shorter than that other of sciaenids. However the weight of sonic muscle did not significantly differ although it was still the lightest one (2.33¡Ó1.00g). The amount of protein (20.37¡Ó0.67¢M) and glycogen (0.33¡Ó0.11¢M) in O. ruber was higher than that of other species and the cross section of sonic muscle fiber was smaller. These characteristics are suitable for O. ruber to make a high-frequency sound. Because of the sound is believed to be made by rubber, so passive sonar was applied to investigate the distribution of this sound type on the estuaries along the west coast of Taiwan (Tam-Shui River, Tou-Chien River, Ta-Chia River, Cho-sui River, Zeng-Wen River, and Kao-Ping River) to represent distribution of O. ruber and its seasonal change of vocal activity. Acoustic activity reached it peak in spring then decreased through summer, autumn, and winter. More sounds were found in the estuaries south of the Cho-sui River (including Cho-sui, Zeng-Wen, and Kao-Ping River) than those north of this river (including Ta-Chia, Tou-Chien, and Tam-Shui River).

Six sciaenid species including Johnius tingi, Johnius sina, Pennahia argentata, Pennahia pawak, Chrysochir aureus and Otolithes rubber from the western coast of Taiwan were studied. Pennahia argentata¡¦s disturbance sound was most distinct in which the pulse period is almost 2~3 times larger than that the other five sciaenid species. The dominant frequencies of C. aureus and O. rubber¡¦s disturbance calls were low (i.e. usually below 500 Hz). In the six sciaenids there was significant difference in the pulse repetition rate, which may be a useful parameter for recognition of sciaenid species. Among the other fishes surveyed in this study, the inter-pulse interval of pearlperch¡¦s (Glaucosoma buergeri) disturbance call is very short or nearly zero. The pulse is composed a low frequency and a high frequency parts, and the frequency range is wide. Most energy was concentrated in the 2nd and 3rd pulses of the Priacanthus macracanthus¡¦s disturbance soundand its pulse period is not stable. The special acoustic character of Pelates quadrilineatus¡¦s disturbance sound is the extremely short inter-pulse interval (< 1ms).The seacatfish Arius maculates¡¦s disturbance sound can be sorted into three types. Type¢¹is a low frequency sound and usually under 1000Hz; it is very similar to sciaenids¡¦ calls with stable pulse period and dominant frequency. Compared to Type¢¹, Type¢º and Type¢» were high-frequency sounds (i.e. usually up to 8kHz) and their pulse periods are less stable. Pomadasys kaakan¡¦s disturbance sound is emitted by stridulating of its pharyngeal teeth and these sounds were high frequency (i.e. usually higher than 10 kHz). Both Johnius tingi females and males possess extrinsic sonic muscles. The dominant frequency of the males¡¦ disturbance call is significant higher than that of females¡¦ and the length of males¡¦ sonic muscles are also significantly longer than that of females¡¦. It is hypothesized that a longer sonic muscle can generate higher tension leading to a higher frequency sound. Comparing my data to the eleven sound types collected from the western coast by other workers in which the sound producers are still unknown, Type A, Type F, and Type G are more similar to that of the disturbance sound of J. sina¡BP. pawak and J. belengerii, respectively.

Johnius specis (Sciaenidae) in Taiwanese waters include five species, Johnius amblycephalus, Johnius belengerii, Johnius macrohynus, Johnius dussumieri and Johnius distinctus. They are soniferous fishes. Contraction and relaxation of the sonic muscles cause the swim bladder to pulsate thus generating and amplifying the sounds. Among these species, only males of J. belengerii and J. macrohynus possess sonic muscle, whereas both sexes of the other three species possess sonic muscles. Proteomic characteristics in the male and female sonic muscles are studied in order to observe if obvious dimorphism exists. CO1-gene sequences were used to reconstruct the phylogenetic tree for these five species. The evolutionary pathway of this tree should suggest the evolutionary change of the sexual dimorphism in the sonic system. Various indices relating to muscle size were higher in male. 17 protein spots were shown in the 2D page for male and female sonic muscles, including 5 structural proteins (Actin alpha skeletal muscle, Tropomyosin-1, Myosin light chain 2, Myosin light chain 3, Myosin light polypeptide 4) and 10 metabolic proteins (Parvalbumin, ATP synthase subunit beta, GTP cyclohydrolase, creatine kinase isoform £\, Muscle-type creatine kinase, Putative pterin-4-alpha-carbinolamine dehydratase, Beta-1,4-galactosyltransferase, Protein-glutamine gamma-glutamyltransferase 4, Triosephosphate isomerase, creatine kinase1 ) and stress protein(Heat shock protein 70) . Differences in expression between the sexes were discussed. The resulted phylogenetic trees show that J. belengerii and J. macrohynus are more closely related than to the other three species, which are more basal and members not belong to a specific clade. Whether the common ancestor of Johnius possessed the sonic muscles remains unclear.

Ophidiidae are major benthopelagic fishes with wide distribution and depth range. Three types of sonic structural patterns have been described in some studied ophidiid fishes, however, the significance of the different types of the sonic apparatus, and the distribution in the subfamilies remain unclear. In addition, although the sonic apparatus and the sonic mechanisms of the high frequency sounds produced by cusk-eels has been explained, the characteristics of extrinsic swimbladder sonic muscle is still unknown. In this study, I investigated three ophidiid subfamilies from Taiwan and reviewed the published data to study the morphological traits of their sonic apparatus. Inaddition, Hoplobrotula armata were related to study the proteomic characterization of the extrinsic swimbladder sonic muscle. Results indicate that the 13 ophidiid species in this study could be grouped into five sonic structural patterns, and three subfamilies could be separated accordingly. Furthermore, because the sister group of the family Ophidiidae-carapidae is soniferous, I suggest that the common ancestor of the three ophidiid subfamilies should also be soniferous. Comparisons made in regarded to environmental factors indicate that diversity of sonic apparatus in three ophidiids subfamiliesis depth dependent. The extrinsic swimbladder sonic muscles could be separated into ventral sonic muscle and intermediate sonic muscle. The histological cross-sections of the ventral sonic muscle fibers show peripheral ring myofibrillar region and central core and they are similar with the sonic muscle. Ventral muscle weights were higher in males than females, but the muscle fibers are smaller in males. Protein values of ventral sonic muscles showed high expression in fast and long duration constructional proteins, and males were higher in protein expression than females. Intermediate sonic muscles, on the other hand, were larger in females than males. The cross-sections of muscle fibers were similar to the white muscle. Expression in the fast constructional related protein in the intermediate sonic muscle was higher than ventral sonic muscles, and the metabolically related protein was lower than ventral sonic muscles. The phylogeny of ophidiiform fishes is not clear at present; the sonic-apparatus diversity in ophidiid fishes observed in this study becomes useful to reveal the relationship phylogenetic of ophidiids fishes. Regard the physiology of sound production, I suggest that the larger ventral sonic muscle in males are be provides a better constructional ability, and their smaller fibers are adaptative for energy metabolism regarded for continuous fast constraction and fatigue resistance. The longer intermediate sonic muscle in females is a better constructional ability than male. In intermediate sonic muscle were be fast constructional related protein, higher in expression than the ventral sonic muscles, the metabolic related protein was lower than the ventral sonic muscles. These results suggest that the intermediate sonic muscle has a hight constructional ability but has a disventage of being easily fatigue.

Some fishes use sound to communicate. The majority of these soniferous fishes use superfast sonic muscles to set the vibration of the swim bladder which results in sound emission and sound amplification. Carapus, a benthic ophiidiform genus, use a slow contracting sonic muscle to pull the anteriormost part of the swim bladder, upon termination of the pulling action, the front part of the swim bladder is snap back setting the swimbladder fenestrum to vibrate. This vibration gives rise to the sound. Other ophiidiform fishes may also use a similar way to emit sounds. Among the soniferous percoids, an advanced perform suborder, glaucosomatid, pempherid, and terapontid share a fenestrum-like structure in the front part of their swim bladder. Previous molecular study suggested that the first two groups form a clade (monophyletic group). It is of great interest to compare the proteomic features of these groups with that of the ophiidiform representatives so that the effects of function and phylogeny to the proteomic characteristics of the sonic muscle can be compared. A species was selected for each of these four groups and their proteomics were analyzed. Results of this study, however, revealed the protein composition of the sonic muscles in the ophiidiform species was more similar to that of the pempherid species. The proteins contribute to the close relationship between these two groups was discussed. A total of 484 protein spots were found in these four species and only five were presented in the sonic muscles of all four species, but absent in the white muscles; and only three of them were successfully identified as: Flotillin-1 (spot 6), HBS1-like protein (spot 8), and Ras-related protein ralB-B (spot 10). Their functions which may be related to the specific role of the sonic muscle were discussed.

Work on sound production of deep-sea fishes has been limited to anatomy, and no sounds from identified species have been recorded on the continental slope. Here I examined the sonic muscles of six species in the family Macrouridae by depth (Coelorhincus carminatus, Nezumia bairdii, Coryphaenoides rupestris, Nezumia equalis, Coryphaenoides armatus, Coryphaenoides carapinus,). Due to increasingly limited food with depth, I hypothesized that sonic muscle development would decrease with depth. Sonic muscles were intrinsic and occurred in males and females. Swimbladder and sonic muscle dimensions increased linearly with fish size, but there were no clear differences with depth suggesting sound production remains important in deeper species.

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