The bottlenose dolphin (Tursiops truncatus): A novel model for studying healthy arterial aging.

Endothelial function declines with aging and independently predicts future cardiovascular disease (CVD) events. Diving also impairs endothelial function in humans. Yet, dolphins, being long-lived mammals adapted to diving, undergo repetitive cycles of tissue hypoxia-reoxygenation and disturbed shear stress without manifesting any apparent detrimental effects, as CVD is essentially nonexistent in these animals. Thus, dolphins may be a unique model of healthy arterial aging and may provide insights into strategies for clinical medicine. Emerging evidence shows that the circulating milieu (bioactive factors in the blood) is at least partially responsible for transducing reductions in age-related endothelial function. To assess if dolphins have preserved endothelial function with aging due to a protected circulating milieu, we tested if the serum (pool of the circulating milieu) of bottlenose dolphins (Tursiops truncatus) induces the same arterial aging phenotype as the serum of age-equivalent humans. We incubated conduit arteries from young and old mice with dolphin and human serum and measured endothelial function ex vivo via endothelium-dependent dilation to acetylcholine. While young arteries incubated with serum from mid-life/older adult human serum had lower endothelial function, those incubated with dolphin serum consistently maintained high endothelial function regardless the age of the donor. Thus, studying the arterial health of dolphins could lead to potential novel therapeutic strategies to improve age-related endothelial dysfunction in humans.

Role of the circulating milieu in age-related arterial dysfunction: a novel ex vivo approach.

The circulating milieu, bioactive molecules in the bloodstream, is altered with aging and interfaces constantly with the vasculature. This anatomic juxtaposition suggests that circulating factors may actively modulate arterial function. Here, we developed a novel, translational experimental model that allows for direct interrogation of the influence of the circulating milieu on age-related arterial dysfunction (aortic stiffening and endothelial dysfunction). To do so, we exposed young and old mouse arteries to serum from young and old mice and young and midlife/older (ML/O) adult humans. We found that old mouse and ML/O adult human, but not young, serum stiffened young mouse aortic rings, assessed via elastic modulus (mouse and human serum, P = 0.003 vs. young serum control), and impaired carotid artery endothelial function, assessed by endothelium-dependent dilation (EDD) (mouse serum, P < 0.001; human serum, P = 0.006 vs. young serum control). Furthermore, young mouse and human, but not old, serum reduced aortic elastic modulus (mouse serum, P = 0.009; human serum, P < 0.001 vs. old/MLO serum control) and improved EDD (mouse and human serum, P = 0.015 vs. old/MLO serum control) in old arteries. In human serum-exposed arteries, in vivo arterial function assessed in the human donors correlated with circulating milieu-modulated arterial function in young mouse arteries (aortic stiffness, r = 0.634, P = 0.005; endothelial function, r = 0.609, P = 0.004) and old mouse arteries (aortic stiffness, r = 0.664, P = 0.001; endothelial function, r = 0.637, P = 0.003). This study establishes novel experimental approaches for directly assessing the effects of the circulating milieu on arterial function and implicates changes in the circulating milieu as a mechanism of in vivo arterial aging.NEW & NOTEWORTHY Changes in the circulating milieu with advancing age may be a mechanism underlying age-related arterial dysfunction. Ex vivo exposure of young mouse arteries to the circulating milieu from old mice or midlife/older adults impairs arterial function whereas exposure of old mouse arteries to the circulating milieu from young mice or young adults improves arterial function. These findings establish that the circulating milieu directly influences arterial function with aging.

The gut microbiome as a modulator of arterial function and age-related arterial dysfunction.

The arterial system is integral to the proper function of all other organs and tissues. Arterial function is impaired with aging, and arterial dysfunction contributes to the development of numerous age-related diseases, including cardiovascular diseases. The gut microbiome has emerged as an important regulator of both normal host physiological function and impairments in function with aging. The purpose of this review is to summarize more recently published literature demonstrating the role of the gut microbiome in supporting normal arterial development and function and in modulating arterial dysfunction with aging in the absence of overt disease. The gut microbiome can be altered due to a variety of exposures, including physiological aging processes. We explore mechanisms by which the gut microbiome may contribute to age-related arterial dysfunction, with a focus on changes in various gut microbiome-related compounds in circulation. In addition, we discuss how modulating circulating levels of these compounds may be a viable therapeutic approach for improving artery function with aging. Finally, we identify and discuss various experimental considerations and research gaps/areas of future research.

Biomarkers related to gas embolism: Gas score, pathology, and gene expression in a gas bubble disease model.

Fish exposed to water supersaturated with dissolved gas experience gas embolism similar to decompression sickness (DCS), known as gas bubble disease (GBD) in fish. GBD has been postulated as an alternative to traditional mammals' models on DCS. Gas embolism can cause mechanical and biochemical damage, generating pathophysiological responses. Increased expression of biomarkers of cell damage such as the heat shock protein (HSP) family, endothelin 1 (ET-1) or intercellular adhesion molecule 1 (ICAM-1) has been observed, being a possible target for further studies of gas embolism. The GBD model consisted of exposing fish to supersaturation in water with approximately 170% total dissolved gas (TDG) for 18 hours, producing severe gas embolism. This diagnosis was confirmed by a complete histopathological exam and the gas score method. HSP70 showed a statistically significant upregulation compared to the control in all the studied organs (p <0.02). Gills and heart showed upregulation of HSP90 with statistical significance (p = 0.015 and p = 0.02, respectively). In addition, HSP70 gene expression in gills was positively correlated with gas score (p = 0.033). These results suggest that gas embolism modify the expression of different biomarkers, with HSP70 being shown as a strong marker of this process. Furthermore, gas score is a useful tool to study the abundance of gas bubbles, although individual variability always remains present. These results support the validity of the GBD model in fish to study gas embolism in diseases such as DCS.

Cetacean Intracytoplasmic Eosinophilic Globules: A Cytomorphological, Histological, Histochemical, Immunohistochemical, and Proteomic Characterization.

The nature, etiopathogenesis, and clinicopathologic relevance of the prevalent intracytoplasmic eosinophilic globules (IEGs) within hepatocytes of cetaceans are unknown. This study aims to evaluate the presence and characterize the IEGs in the hepatocytes of cetaceans using histochemical and immunohistochemical electron microscopy, Western blot, lectin histochemistry, and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry techniques. A total of 95/115 (83%) animals (16 species) exhibited histologically evident intracytoplasmic round to oval, single to multiple, hyaline eosinophilic globules within the hepatocytes. These globules were largely PAS-positive, diastase resistant, and were immunopositive for fibrinogen (FB, 97%), albumin (Alb, 85%), and α1-antitrypsine (A1AT, 53%). The IEG positivity for FB and A1AT were correlated with live-stranding, hepatic congestion and a good nutritional status. The cetaceans lacking IEGs were consistently dead stranded and had poor body conditions. The IEGs in 36 bycaught cetaceans were, all except one, FB-positive and A1AT-negative. The IEGs exhibited morphologic and compositional variations at the ultrastructural level, suggesting various stages of development and/or etiopathogenesis(es). The glycocalyx analysis suggested an FB- and A1AT-glycosylation pattern variability between cetaceans and other animals. The proteomic analyses confirmed an association between the IEGs and acute phase proteins, suggesting a relationship between acute stress (i.e., bycatch), disease, and cellular protective mechanisms, allowing pathologists to correlate this morphological change using the acute hepatocytic cell response under certain stress conditions.

Morphological Structure of the Aortic Wall in Deep Diving Cetacean Species: Evidence for Diving Adaptation.

This study analyses the aortic wall structure in nine cetacean species with deep diving habits belonging to four Odontoceti families: Ziphiidae, Kogiidae, Physteridae, and Delphinidae. Samples of ascending, thoracic and abdominal aorta were processed for histological and morphometric studies. The elastic component was higher in the proximal aortic segments, and the muscular elements increased distally in all cases. Morphometric analyses showed that all families presented a decrease in the thickness of the arterial wall and the tunica media along the aorta. The reduction was dramatic between ascending and thoracic aorta in the Physeteridae specimens; meanwhile, the other three families showed a more uniform decrease between the ascending, thoracic and abdominal aorta. The decline was not correlated with a reduced elastic or lamellar unit thickness but with a loss of lamellar units. The organization of the elements in the aortic wall did not show essential modifications between the four families, resembling the structure described previously in the shallow and intermediate diving dolphins. Our findings support that the difference in the morphometric characteristics of the different segments in the aortic wall is likely related to the diving habit more than the absolutes values of any other parameter.

Cetaceans Humerus Radiodensity by CT: A Useful Technique Differentiating between Species, Ecophysiology, and Age.

Cetaceans are mammals that underwent a series of evolutionary adaptations to live in the aquatic environment, including morphological modifications of various anatomical structures of the skeleton and their bone mineral density (BMD); there are few studies on the latter. BMD is related to the radiodensity measured through computed tomography (CT) in Hounsfield units (HU). This work aimed to test and validate the usefulness of studying humeral bone radiodensity by CT of two cetacean species (the Atlantic spotted dolphin and the pygmy sperm whale) with different swimming and diving habits. The radiodensity was analysed at certain levels following a new protocol based on a review of previous studies. Humeral radiodensity values were related to four aspects: species, diving behaviour, swimming activity level, and age. We observed that the consistent differences in the radiodensity of the cortical bone of the distal epiphysis between animals of different life-history categories suggest that this bone portion could be particularly useful for future ontogenetic studies. Hence, this technique may be helpful in studying and comparing species with different ecophysiologies, particularly distinguishing between swimming and diving habits.

Establishment of a fish model to study gas-bubble lesions.

Decompression sickness (DCS) is a clinical syndrome caused by the formation of systemic intravascular and extravascular gas bubbles. The presence of these bubbles in blood vessels is known as gas embolism. DCS has been described in humans and animals such as sea turtles and cetaceans. To delve deeper into DCS, experimental models in terrestrial mammals subjected to compression/decompression in a hyperbaric chamber have been used. Fish can suffer from gas bubble disease (GBD), characterized by the formation of intravascular and extravascular systemic gas bubbles, similarly to that observed in DCS. Given these similarities and the fact that fish develop this disease naturally in supersaturated water, they could be used as an alternative experimental model for the study of the pathophysiological aspect of gas bubbles. The objective of this study was to obtain a reproducible model for GBD in fish by an engineering system and a complete pathological study, validating this model for the study of the physiopathology of gas related lesions in DCS. A massive and severe GBD was achieved by exposing the fish for 18 h to TDG values of 162-163%, characterized by the presence of severe hemorrhages and the visualization of massive quantities of macroscopic and microscopic gas bubbles, systemically distributed, circulating through different large vessels of experimental fish. These pathological findings were the same as those described in small mammals for the study of explosive DCS by hyperbaric chamber, validating the translational usefulness of this first fish model to study the gas-bubbles lesions associated to DCS from a pathological standpoint.

Pulmonary and Systemic Skeletal Muscle Embolism in a Beaked Whale with a Massive Trauma of Unknown Aetiology.

An adult female Sowerby's beaked whale was found floating dead in Hermigua (La Gomera, Canary Islands, Spain) on 7 December 2016. Severe traumas of unknown aetiology were attributed, and the gross and microscopic findings are consistent with catastrophic trauma as a cause of death. Rib fractures affected the intercostals, transverse thoracis skeletal muscles, and thoracic rete mirabile. Degenerated muscle fibres were extruded to flow into vascular and lymphatic vessels travelling to several anatomic locations into the thoracic cavity, including the lungs, where they occluded the small lumen of pulmonary microvasculature. A pulmonary and systemic skeletal muscle embolism was diagnosed, constituting the first description of this kind of embolism in an animal. The only previous description has been reported in a woman after peritoneal dialysis. Skeletal pulmonary embolism should be considered a valuable diagnostic for different types of trauma in vivo in wild animals. This is especially valuable when working with decomposed carcasses, as in those cases, it is not always feasible to assess other traumatic evidence.

Decompressive Pathology in Cetaceans Based on an Experimental Pathological Model.

Decompression sickness (DCS) is a widely known clinical syndrome in human medicine, mainly in divers, related to the formation of intravascular and extravascular gas bubbles. Gas embolism and decompression-like sickness have also been described in wild animals, such as cetaceans. It was hypothesized that adaptations to the marine environment protected them from DCS, but in 2003, decompression-like sickness was described for the first time in beaked whales, challenging this dogma. Since then, several episodes of mass strandings of beaked whales coincidental in time and space with naval maneuvers have been recorded and diagnosed with DCS. The diagnosis of human DCS is based on the presence of clinical symptoms and the detection of gas embolism by ultrasound, but in cetaceans, the diagnosis is limited to forensic investigations. For this reason, it is necessary to resort to experimental animal models to support the pathological diagnosis of DCS in cetaceans. The objective of this study is to validate the pathological results of cetaceans through an experimental rabbit model wherein a complete and detailed histopathological analysis was performed. Gross and histopathological results were very similar in the experimental animal model compared to stranded cetaceans with DCS, with the presence of gas embolism systemically distributed as well as emphysema and hemorrhages as primary lesions in different organs. The experimental data reinforces the pathological findings found in cetaceans with DCS as well as the hypothesis that individuality plays an essential role in DCS, as it has previously been proposed in animal models and human diving medicine.

Myoglobin Concentration and Oxygen Stores in Different Functional Muscle Groups from Three Small Cetacean Species.

Compared with terrestrial mammals, marine mammals possess increased muscle myoglobin concentrations (Mb concentration, g Mb · 100g-1 muscle), enhancing their onboard oxygen (O2) stores and their aerobic dive limit. Although myoglobin is not homogeneously distributed, cetacean muscle O2 stores have been often determined by measuring Mb concentration from a single muscle sample (longissimus dorsi) and multiplying that value by the animal's locomotor muscle or total muscle mass. This study serves to determine the accuracy of previous cetacean muscle O2 stores calculations. For that, body muscles from three delphinid species: Delphinus delphis, Stenella coeruleoalba, and Stenella frontalis, were dissected and weighed. Mb concentration was calculated from six muscles/muscle groups (epaxial, hypaxial and rectus abdominis; mastohumeralis; sternohyoideus; and dorsal scalenus), each representative of different functional groups (locomotion powering swimming, pectoral fin movement, feeding and respiration, respectively). Results demonstrated that the Mb concentration was heterogeneously distributed, being significantly higher in locomotor muscles. Locomotor muscles were the major contributors to total muscle O2 stores (mean 92.8%) due to their high Mb concentration and large muscle masses. Compared to this method, previous studies assuming homogenous Mb concentration distribution likely underestimated total muscle O2 stores by 10% when only considering locomotor muscles and overestimated them by 13% when total muscle mass was considered.

Retrospective Study of Fishery Interactions in Stranded Cetaceans, Canary Islands.

Estimating cetacean interactions with fishery activities is challenging. Bycatch and chronic entanglements are responsible for thousands of cetacean deaths per year globally. This study represents the first systematic approach to the postmortem investigation of fishery interactions in stranded cetaceans in the Canary Islands. We retrospectively studied 586 cases necropsied between January 2000 and December 2018. Of the cases with a known cause of death, 7.4% (32/453) were due to fishery interactions, and the Atlantic spotted dolphin (Stenella frontalis) was the most affected species [46.9% (15/32)]. Three types of fishery interactions were recognized by gross findings: bycatch [65.6% (21/32)], chronic entanglements [18.8% (6/32)], and fishermen aggression [15.6% (5/32)]. Among the bycaught cases, we differentiated the dolphins that died because of ingestion of longline hooks [23.8% (5/21)] from those that died because of fishing net entrapments [76.2% (16/21)], including dolphins that presumably died at depth due to peracute underwater entrapment (PUE) [37.5% (6/16)], dolphins that were hauled out alive and suffered additional trauma during handling [43.8% (7/16)], and those that were released alive but became stranded and died because of fishery interactions [18.7% (3/16)]. Gross and histologic findings of animals in each group were presented and compared. The histological approach confirmed gross lesions and excluded other possible causes of death. Cetaceans in good-fair body condition and shallow diving species were significantly more affected by fishery interactions, in agreement with the literature. Low rates of fishery interactions have been described, compared with other regions. However, within the last few years, sightings of entangled live whales, especially the minke whale (Balaenoptera acutorostrata) and Bryde's whale (B. edeni), have increased. This study contributes to further improvement of the evaluation of different types of fishery interactions and may facilitate the enforcement of future conservation policies to preserve cetacean populations in the Canary Islands.

Amyloid-beta peptide and phosphorylated tau in the frontopolar cerebral cortex and in the cerebellum of toothed whales: aging versus hypoxia.

Hypoxia could be a possible risk factor for neurodegenerative alterations in cetaceans' brain. Among toothed whales, the beaked whales are particularly cryptic and routinely dive deeper than 1000 m for about 1 h in order to hunt squids and fishes. Samples of frontal cerebral and cerebellar cortex were collected from nine animals, representing six different species of the suborder Odontoceti. Immunohistochemical analysis employed anti-ß-amyloid (Aß) and anti-neurofibrillary tangle (NFT) antibodies. Six of nine (67%) animals showed positive immunolabeling for Aß and/or NFT. The most striking findings were intranuclear Aß immunopositivity in cerebral cortical neurons and NFT immunopositivity in cerebellar Purkinje neurons with granulovacuolar degeneration. Aß plaques were also observed in one elderly animal. Herein, we present immunohistopathological findings classic of Alzheimer's and other neurodegenerative diseases in humans. Our findings could be linked to hypoxic phenomena, as they were more extensive in beaked whales. Despite their adaptations, cetaceans could be vulnerable to sustained and repetitive brain hypoxia.

Lipids of lung and lung fat emboli of the toothed whales (Odontoceti).

Lipids are biomolecules present in all living organisms that, apart from their physiological functions, can be involved in different pathologies. One of these pathologies is fat embolism, which has been described histologically in the lung of cetaceans in association with ship strikes and with gas and fat embolic syndrome. To assess pathological lung lipid composition, previous knowledge of healthy lung tissue lipid composition is essential; however, these studies are extremely scarce in cetaceans. In the present study we aimed first, to characterize the lipids ordinarily present in the lung tissue of seven cetacean species; and second, to better understand the etiopathogenesis of fat embolism by comparing the lipid composition of lungs positive for fat emboli, and those negative for emboli in Physeter macrocephalus and Ziphius cavirostris (two species in which fat emboli have been described). Results showed that lipid content and lipid classes did not differ among species or diving profiles. In contrast, fatty acid composition was significantly different between species, with C16:0 and C18:1ω9 explaining most of the differences. This baseline knowledge of healthy lung tissue lipid composition will be extremely useful in future studies assessing lung pathologies involving lipids. Concerning fat embolism, non-significant differences could be established between lipid content, lipid classes, and fatty acid composition. However, an unidentified peak was only found in the chromatogram for the two struck whales and merits further investigation.

Retrospective Study of Traumatic Intra-Interspecific Interactions in Stranded Cetaceans, Canary Islands.

Aggressive encounters involving cetacean species are widely described in the literature. However, detailed pathological studies regarding lesions produced by these encounters are scarce. From January 2000 to December 2017, 540 cetaceans stranded and were necropsied in the Canary Islands, Spain. Of them, 24 cases of eight species presented social traumatic lesions produced by cetaceans of the same or different species. All the cases presented severe multifocal vascular changes, 50% (12/24) presented fractures affecting mainly the thoracic region, 41.7% (10/24) acute tooth-rake marks, 37.5% (9/24) undigested food in the stomach, 33.3% (8/24) tracheal edema, and 12.5% (3/24) pulmonary perforation. In 10 cases with tooth-rake marks, the distance between the teeth, allowed us to further identify the aggressor species: four cases were compatible with killer whales (Orcinus orca) affecting three species [pigmy sperm whale (Kogia breviceps), Cuvier's beaked whale (Ziphius cavirostris), and short-finned pilot whale (Globicephala macrorhynchus)] and four cases compatible with common bottlenose dolphins (Tursiops truncatus) affecting two species [short-beaked common dolphin (Delphinus delphis) and Atlantic spotted dolphin (Stenella frontalis)]. We also described two cases of intraspecific interaction in stripped dolphin (Stenella coeruleoalba). Microscopically, 70.8% (17/24) of the cases presented acute degenerative myonecrosis, 66.7% (14/21) presented vacuoles in the myocardiocytes, 36.8% (7/19) pigmentary tubulonephrosis, 31.6% (6/19) cytoplasmic eosinophilic globules within hepatocytes, 21.4% (3/14) hemorrhages in the adrenal gland, and 17.3% (4/23) bronchiolar sphincter contraction. The statistical analysis revealed that deep divers, in good body condition and nearby La Gomera and Tenerife were more prone to these fatal interactions. Additionally, in this period, three animals died due to an accident during predation: a false killer whale (Pseudorca crassidens) died because of a fatal attempt of predation on a stingray, and two Risso's dolphins (Grampus griseus) died as a consequence of struggling while predating on large squids.

Morphological structure of the aortic wall in three Delphinid species with shallow or intermediate diving habits: Evidence for diving adaptation.

Some modifications in the vascular system of marine mammals provide adaptive advantages for diving. This study analyses the organisation of the aortic wall in dolphins, observing artery changes in volume and blood pressure for diving behaviour. Samples of three aortic segments (ascending, thoracic and abdominal) of three dolphin species were processed for histological and morphometric studies. The three dolphin species used, striped dolphin (Stenella coeruleoalba), Atlantic spotted dolphin (Stenella frontalis) and common dolphin (Delphinus delphis), have shallow or intermediate diving habits. Our results indicated that the components of the aortic wall of the dolphins had different dispositions in the three selected segments. The aortic wall decreased in thickness along its length due to a loss of the lamellar units in the tunica media and a thinning of the main elements of the lamellar units along the artery. The life stage had little influence on the thickness of the aortic wall except for the ascending aorta. The weight, body length, species or sex of the specimen did not significantly influence the thickness of the wall or the lamellar units. In summary, the histological and morphometric aortic structure in dolphins, in relation to the studied parameters, seems to be similar to that previously described of terrestrial mammals such as pigs, except for a larger difference in the proportion of lamellar units between the ascending and thoracic segments.

Capture Myopathy and Stress Cardiomyopathy in a Live-Stranded Risso's Dolphin (Grampus griseus) in Rehabilitation.

Capture myopathy (CM) is described in wild animals as a metabolic syndrome resulting from the extreme stress suffered during and after capture, handling, restraint, and transport. Although CM has been characterized in many species of cetaceans, descriptions of cardiac injury-an important component of this syndrome, and, according to previous authors, comparable to the existing human pathology so-called stress cardiomyopathy (SCMP)-are still rare. Therefore, the main aim of this report is to illustrate, for the first time, the biochemical analysis, and gross, histopathological, histochemical and immunohistochemical features of CM, and more specifically of the SCMP involved in this syndrome, caused by the live-stranding and consequent rehabilitation attempt, for a certain period of time, in a juvenile male Risso's dolphin (Grampus griseus). The animal presented elevated values of creatine kinase, cardiac troponin I and blood urea nitrogen, with some variations during the rehabilitation period. Histologically, we detected vascular changes and acute degenerative lesions analogous to the ones observed in humans with SCMP. We consider this study to be an important contribution to the study of cetaceans since it could help in decision-making and treatment procedures during live-strandings and improve conservation efforts by reducing the mortality of these animals.

Comparative morphology, histology, and cytology of odontocete cetaceans prostates.

The prostate is the only male accessory gland in cetaceans. However, little is known about this organ in these species. Anatomical and histological characteristics of the prostate have been described in only a few cetacean species, further, one study reported a high incidence of prostatic pathologies in cetaceans that may impair reproduction. The objective of this work was to describe and compare the morphological, histological, and cytological characteristics of the prostate in different odontocete cetaceans. To this end, the prostate glands of 47 animals from nine different species of cetaceans were macroscopically and microscopically studied. Members of the families Delphinidae, Ziphiidae, and Physeteridae were included. In general, the prostate appeared as a musculo-glandular organ with two distinct parts-the Corpus prostatae and the Pars disseminata prostatae. In the pygmy sperm whale (Kogia breviceps) and the Cuvier's beaked whale (Ziphius cavirostris), the prostate was a discrete gland with a small Corpus prostatae. Microscopically, the prostates of different delphinids species shared similarities; however, the prostate of the pygmy sperm whale revealed significant histological differences compared to those of the delphinids. Immunohistochemical analysis was performed using low- and high-molecular-weight cytokeratin, vimentin, and prostatic specific antigen commercial antibodies. Electron microscopy analysis was performed on the prostate of a bottlenose dolphin and the cytomorphological differences among the major epithelial components of the prostatic epithelium were described. Anat Rec, 2019. © 2019 American Association for Anatomy Anat Rec, 303:2036-2053, 2020. © 2019 American Association for Anatomy.

Locus coeruleus complex of the family Delphinidae.

The locus coeruleus (LC) is the largest catecholaminergic nucleus and extensively projects to widespread areas of the brain and spinal cord. The LC is the largest source of noradrenaline in the brain. To date, the only examined Delphinidae species for the LC has been a bottlenose dolphin (Tursiops truncatus). In our experimental series including different Delphinidae species, the LC was composed of five subdivisions: A6d, A6v, A7, A5, and A4. The examined animals had the A4 subdivision, which had not been previously described in the only Delphinidae in which this nucleus was investigated. Moreover, the neurons had a large amount of neuromelanin in the interior of their perikarya, making this nucleus highly similar to that of humans and non-human primates. This report also presents the first description of neuromelanin in the cetaceans' LC complex, as well as in the cetaceans' brain.