Perineal muscles and motoneurons are sexually monomorphic in the naked mole‐rat (Heterocephalus glaber)

Naked mole‐rats are eusocial mammals that live in colonies with a single breeding female and one to three breeding males. All other members of the colony, known as subordinates, are nonreproductive and exhibit few sex differences in behavior or genital anatomy. This raises questions about the degree of sexual differentiation in subordinate naked mole‐rats. The striated perineal muscles associated with the phallus [the bulbocavernosus (BC), ischiocavernosus (IC), and levator ani (LA) muscles], and their innervating motoneurons, are sexually dimorphic in all rodents examined to date. We therefore asked whether perineal muscles and motoneurons were also sexually dimorphic in subordinate naked mole‐rats. Muscles similar to the LA and IC of other rodents were found in naked mole‐rats of both sexes. No clear BC muscle was identified, although a large striated muscle associated with the urethra in male and female naked mole‐rats may be homologous to the BC of other rodents. There were no sex differences in the volumes of the LA, IC, or the urethral muscles. Motoneurons innervating the perineal muscles were identified by retrograde labeling with cholera‐toxin‐conjugated horseradish peroxidase. All perineal motoneurons were found in a single cluster in the ventrolateral lateral horn, in a position similar to that of Onuf's nucleus of carnivores and primates. There was no sex difference in the size or number of motoneurons in Onuf's nucleus of naked mole‐rats. Thus, unlike findings in any other mammal, neither the perineal muscles nor the perineal motoneurons appear to be sexually differentiated in subordinate naked mole‐rats. © 2002 Wiley Periodicals, Inc. J Neurobiol 51: 33–42, 2002     

Genetic diversity of the naked mole‐rat (Heterocephalus glaber)

The naked mole‐rat (Heterocephalus glaber) is used as an animal model in various studies, but not much is known on the genetic diversity of this animal. Here, on the basis of dataset collected from the most part of the distribution range of the naked mole‐rat, we reconstruct phylogenetic relationships between its different lineages using mitochondrial and nuclear markers. We also mapped the distribution of the main genetic lineages, dated the divergence using different Bayesian tree‐calibration techniques, and modeled the distribution of ecological niches for the period of last glacial maximum. Our results show the existence of two deeply divergent clades designated as the eastern clade (East Ethiopia) and the southern clade (South Ethiopia and North Kenya). Additional phylogeographic structure was demonstrated for each of these two clades. Divergence between these two main lineages dated back to the Middle Pleistocene (ca. 1.4–0.8 Mya) and may have been related to climate changes in Africa during the Mid‐Pleistocene Revolution. In light of substantial genetic differences between the eastern and southern lineages of the naked mole‐rat, these two clades can be considered as two deeply divergent subspecies or even as distinct species.     

Challenging the inbreeding hypothesis in a eusocial mammal: population genetics of the naked mole‐rat,Heterocephalus glaber

The role of genetic relatedness in the evolution of eusociality has been the topic of much debate, especially when contrasting eusocial insects with vertebrates displaying reproductive altruism. The naked mole‐rat, Heterocephalus glaber, was the first described eusocial mammal. Although this discovery was based on an ecological constraints model of eusocial evolution, early genetic studies reported high levels of relatedness in naked mole‐rats, providing a compelling argument that low dispersal rates and consanguineous mating (inbreeding as a mating system) are the driving forces for the evolution of this eusocial species. One caveat to accepting this long‐held view is that the original genetic studies were based on limited sampling from the species’ geographic distribution. A growing body of evidence supports a contrary view, with the original samples not representative of the species—rather reflecting a single founder event, establishing a small population south of the Athi River. Our study is the first to address these competing hypotheses by examining patterns of molecular variation in colonies sampled from north and south of the Athi and Tana rivers, which based on our results, serve to isolate genetically distinct populations of naked mole‐rats. Although colonies south of the Athi River share a single mtDNA haplotype and are fixed at most microsatellite loci, populations north of the Athi River are considerably more variable. Our findings support the position that the low variation observed in naked mole‐rat populations south of the Athi River reflects a founder event, rather than a consequence of this species’ unusual mating system.     

Metabolic clues to salubrious longevity in the brain of the longest‐lived rodent: the naked mole‐rat

Naked mole‐rats (NMRs) are the oldest‐living rodent species. Living underground in a thermally stable ecological niche, NMRs have evolved certain exceptional traits, resulting in sustained health spans, negligible cognitive decline, and a pronounced resistance to age‐related disease. Uncovering insights into mechanisms underlying these extraordinary traits involved in successful aging may conceivably provide crucial clues to extend the human life span and health span. One of the most fundamental processes inside the cell is the production of ATP, which is an essential fuel in driving all other energy‐requiring cellular activities. Not surprisingly, a prominent hallmark in age‐related diseases, such as neurodegeneration and cancer, is the impairment and dysregulation of metabolic pathways. Using a two‐dimensional polyacrylamide gel electrophoresis proteomics approach, alterations in expression and phosphorylation levels of metabolic proteins in the brains of NMRs, aged 2–24 years, were evaluated in an age‐dependent manner. We identified 13 proteins with altered levels and/or phosphorylation states that play key roles in various metabolic pathways including glycolysis, β‐oxidation, the malate‐aspartate shuttle, the Tricarboxylic Acid Cycle (TCA) cycle, the electron transport chain, NADPH production, as well as the production of glutamate. New insights into potential pathways involved in metabolic aspects of successful aging have been obtained by the identification of key proteins through which the NMR brain responds and adapts to the aging process and how the NMR brain adapted to resist age‐related degeneration.

     

Functional Morphology of the Olfactory Mucosa and Olfactory Bulb in Fossorial Rodents: The East African Root Rat (Tachyoryctes splendens) and the Naked Mole Rat (Heterocephalus glaber)

Optimal functioning of the olfactory system is critical for survival of fossorial rodents in their subterranean lifestyle. This study examines the structure of the olfactory mucosa and olfactory bulb of two fossorial rodents exhibiting distinct social behaviors, the East African root rat and the naked mole rat. The social naked mole rat displayed simpler ethmoturbinates consisting of dorsomedial and broad discoid/flaplike parts that projected rostrally from the ethmoid bone. In the solitary root rat however, the ethmoturbinates were highly complex and exhibited elaborate branching which greatly increased the olfactory surface area. In addition, when correlated with the whole brain, the volume of the olfactory bulbs was greater in the root rat (4.24 × 10^?2) than in the naked mole rat (3.92 × 10^?2). Results of this study suggest that the olfactory system of the root rat is better specialized than that of the naked mole rat indicating a higher level of dependence on this system since it leads a solitary life. The naked mole rat to the contrary may have compensated for its relatively inferior olfactory system by living in groups in a social system. These findings demonstrate that structure of the olfactory system of fossorial mammals is dictated by both behavior and habitat.     

Kidney concentrating ability of a subterranean xeric rodent,the naked mole-rat (Heterocephalus glaber)

The naked mole-rat (Heterocephalus glaber) is a strictly subterranean mammal inhabiting the arid zones of north-east Africa. These animals have no access to free water and water balance thus might be facilitated by regulating renal water loss. The urinary concentrating ability of the naked mole-rat was determined using five dietary manipulations in which both water and salt content were altered. Control animals (n=7) received a high quality protein cereal mixed to a thin paste with water (1 g cereal: 85 g water). Water stress was induced by reducing the water content of the diet by either 50% (n=7) or 65% (n=7). Salt loading was facilitated by replacing the water with the same volume of either 0.9% salt (n=7) or 3.0% salt (n=4) solutions. Changes in body mass, food consumption and urine volume were measured daily. The effect of diet on osmolality and electrolyte concentrations of urine and plasma were determined on termination of the diet trials. Although energy intake was not reduced, naked mole-rats lost body weight with both water stress treatments. Urine volume voided per day decreased significantly with both water stress treatments (P<0.05), such that the most extreme water stress led to an 80% reduction in urine volume. Mildly salt-loaded animals gained weight, yet underwent a sodium diuresis, as indicated by a 1.3-fold increase in the daily volume of urine voided (P<0.05). Maximum urine concentration (1521±250 mmol·kg^-1) was achieved with mild water stress and was 4.6±0.9 times that of plasma. Neither further water stress nor salt loading further increased urine osmolality (P>0.05). The naked mole-rat exhibits a moderate kidney concentrating ability and cannot maintain plasma osmolality or body mass with either extreme water stress or salt loading. Although this species succesfully inhabits arid zones, survival in these areas is not facilitated by renal water conservation, but rather by their underground existence in a microhabitat where humidities are high and radiant heat loads low. In this milieu a moderate kidney concentrating ability is adequate.Abbreviations Bm body mass - ESL extreme salt load - EWS extreme water stress - MSL mild salt load - MWS mild water stress     

Middle ear structure and bone conduction in Spalax,Eospalax, and Tachyoryctes mole‐rats (Rodentia: Spalacidae)

There is evidence that spalacine, tachyoryctine, and myospalacine mole‐rats all communicate with conspecifics through a form of seismic signaling, but the route for the detection of these signals is disputed. It has been proposed that two unusual anatomical adaptations in Spalax allow jaw vibrations to pass to the inner ear via the incus and stapes: a pseudoglenoid (=postglenoid) fossa which accomodates the condylar process of the mandible, and a bony cup, supported by a periotic lamina, through which the incus articulates with the skull. In this study, a combination of dissection and computed tomography was used to examine the ear region in more detail in both Spalax and its subterranean relatives Tachyoryctes and Eospalax, about which much less is known. Tachyoryctes was found to lack a pseudoglenoid fossa, while Eospalax lacks a periotic lamina and bony cup. This shows that these structures need not simultaneously be present for the detection of ground vibrations in mole‐rats. Based on the observed anatomy, three hypothetical modes of bone conduction are argued to represent more likely mechanisms through which mole‐rats can detect ground vibrations: ossicular inertial bone conduction, a pathway involving sound radiation into the external auditory meatus, and a newly‐described fluid pathway between pseudoglenoid fossa and cranial cavity. The caudolateral extension of the tympanic cavity and the presence of a bony cup might represent synapomorphies uniting Spalax and Tachyoryctes, while the loss of the tensor tympani muscle in Spalax and Eospalax may be convergently derived. J. Morphol., 2010. © 2009 Wiley‐Liss, Inc.     

Parasite‐associated mortality in a long‐lived mammal: Variation with host age,sex, and reproduction

Parasites can cause severe host morbidity and threaten survival. As parasites are generally aggregated within certain host demographics, they are likely to affect a small proportion of the entire population, with specific hosts being at particular risk. However, little is known as to whether increased host mortality from parasitic causes is experienced by specific host demographics. Outside of theoretical studies, there is a paucity of literature concerning dynamics of parasite‐associated host mortality. Empirical evidence mainly focuses on short‐lived hosts or model systems, with data lacking from long‐lived wild or semi‐wild vertebrate populations. We investigated parasite‐associated mortality utilizing a multigenerational database of mortality, health, and reproductive data for over 4,000 semi‐captive timber elephants (Elephas maximus), with known causes of death for mortality events. We determined variation in mortality according to a number of host traits that are commonly associated with variation in parasitism within mammals: age, sex, and reproductive investment in females. We found that potentially parasite‐associated mortality varied significantly across elephant ages, with individuals at extremes of lifespan (young and old) at highest risk. Mortality probability was significantly higher for males across all ages. Female reproducers experienced a lower probability of potentially parasite‐associated mortality than females who did not reproduce at any investigated time frame. Our results demonstrate increased potentially parasite‐associated mortality within particular demographic groups. These groups (males, juveniles, elderly adults) have been identified in other studies as susceptible to parasitism, stressing the need for further work investigating links between infection and mortality. Furthermore, we show variation between reproductive and non‐reproductive females, with mothers being less at risk of potentially parasite mortality than nonreproducers.     

The comparative anatomy of the abdominal gastrointestinal tract of six species of African mole‐rats (Rodentia,Bathyergidae)

The gastrointestinal tracts (GITs) of six species of African mole‐rats (Bathyergidae) were compared. The aim was to provide a comprehensive anatomical comparison between the different species. The relative shape, length, and surface areas were taken into account to determine whether the GITs are phylogenetically constrained or exhibit anatomical adaptations in response to diets. In all six species the stomach was simple and glandular. With the exception of Heterocephalus glaber, the caecum was coiled in a flat spiral, the ascending colon was arranged in a loop of varying lengths, and a mucosal colonic papillary‐lined groove was present in the ascending colon in all species. By contrast, the caecum in H. glaber was uncoiled, the ascending colon was not looped, and the groove was not papillated. A caeco‐appendix was observed only in Bathyergus suillus and Georychus capensis. Hierarchical multivariate cluster analysis on the presence/absence of nine anatomical structures associated with the GIT of mole‐rats revealed that H. glaber was anatomically the least similar of the six species (77.6% similarity) with respect to the nine GIT variables included. All Cryptomys species were the same (100% similarity), and two species B. suillus and G. capensis grouped together and were more similar to the Cryptomys genus (95% similarity) than they were to H. glaber. These findings support previous phylogenetic classifications. The voluminous caeco‐colon in B. suillus may be explained by its ingestion of grasses in addition to below‐ground storage organs of plants. We conclude that phylogeny and diet affect the GIT anatomy of the African mole rats studied here. J. Morphol., 2010. © 2009 Wiley‐Liss, Inc.     

Parentage analysis of Ansell's mole‐rat family groups indicates a high reproductive skew despite relatively relaxed ecological constraints on dispersal

To better understand evolutionary pathways leading to eusociality, interspecific comparisons are needed, which would use a common axis, such as that of reproductive skew, to array species. African mole‐rats (Bathyergidae, Rodentia) provide an outstanding model of social evolution because of a wide range of social organizations within a single family; however, their reproductive skew is difficult to estimate, due to their cryptic lifestyle. A maximum skew could theoretically be reached in groups where reproduction is monopolized by a stable breeding pair, but the value could be decreased by breeding‐male and breeding‐female turnover, shared reproduction and extra‐group mating. The frequency of such events should be higher in species or populations inhabiting mesic environments with relaxed ecological constraints on dispersal. To test this prediction, we studied patterns of parentage and relatedness within 16 groups of Ansell's mole‐rat (Fukomys anselli) in mesic miombo woodland. Contrary to expectation, there was no shared reproduction (more than one breeder of a particular sex) within the studied groups, and proportion of immigrants and offspring not assigned to current breeding males was low. The within‐group parentage and relatedness patterns observed resemble arid populations of ‘eusocial’ Fukomys damarensis, rather than a mesic population of ‘social’ Cryptomys hottentotus. As a possible explanation, we propose that the extent ecological conditions affect reproductive skew may be markedly affected by life history and natural history traits of the particular species and genera.     

Age estimation in a long‐lived seabird (Ardenna tenuirostris) using DNA methylation‐based biomarkers

Age structure is a fundamental aspect of animal population biology. Age is strongly related to individual physiological condition, reproductive potential and mortality rate. Currently, there are no robust molecular methods for age estimation in birds. Instead, individuals must be ringed as chicks to establish known‐age populations, which is a labour‐intensive and expensive process. The estimation of chronological age using DNA methylation (DNAm) is emerging as a robust approach in mammals including humans, mice and some non‐model species. Here, we quantified DNAm in whole blood samples from a total of 71 known‐age Short‐tailed shearwaters (Ardenna tenuirostris) using digital restriction enzyme analysis of methylation (DREAM). The DREAM method measures DNAm levels at thousands of CpG dinucleotides throughout the genome. We identified seven CpG sites with DNAm levels that correlated with age. A model based on these relationships estimated age with a mean difference of 2.8 years to known age, based on validation estimates from models created by repeated sampling of training and validation data subsets. Longitudinal observation of individuals re‐sampled over 1 or 2 years generally showed an increase in estimated age (6/7 cases). For the first time, we have shown that epigenetic changes with age can be detected in a wild bird. This approach should be of broad interest to researchers studying age biomarkers in non‐model species and will allow identification of markers that can be assessed using targeted techniques for accurate age estimation in large population studies.     

Haematological parameters do senesce in the wild: evidence from different populations of a long‐lived mammal

Increasing evidence of senescence has been reported from long‐term studies of wild populations. However, most studies have focused on life‐history traits like survival, reproduction or body mass, generally from a single intensively monitored population. However, variation in the intensity of senescence across populations, and to a lesser extent between sexes, is still poorly understood. In addition, the pattern of age‐specific changes in haematological parameters remains virtually unknown to date for any population of vertebrate living in the wild. Using repeated blood samples collected from known‐aged (2–15 years of age) roe deer (Capreolus capreolus) from two populations facing highly different environmental conditions, we filled the gap. In particular, we investigated age‐specific changes in haematocrit, albumin and creatinine. We reported clear evidence of senescence in all haematological parameters. Moreover, senescence patterns differed between sexes and populations. The rate of senescence was higher in males than in females for haematocrit with no site difference. On the other hand, the rate of senescence in creatinine was higher at Trois Fontaines than at Chizé with no sex difference. Our findings provide a first demonstration of age‐specific declines in haematological parameters in wild populations of large herbivores and show that the process of senescence in vertebrates is not restricted to body mass or fitness components. We also demonstrate that the senescence pattern of haematological parameters is context dependent and varies both between sexes and according to environmental conditions.     

Downregulation of the inflammatory network in senescent fibroblasts and aging tissues of the long‐lived and cancer‐resistant subterranean wild rodent,Spalax

The blind mole rat (Spalax) is a wild, long‐lived rodent that has evolved mechanisms to tolerate hypoxia and resist cancer. Previously, we demonstrated high DNA repair capacity and low DNA damage in Spalax fibroblasts following genotoxic stress compared with rats. Since the acquisition of senescence‐associated secretory phenotype (SASP) is a consequence of persistent DNA damage, we investigated whether cellular senescence in Spalax is accompanied by an inflammatory response. Spalax fibroblasts undergo replicative senescence (RS) and etoposide‐induced senescence (EIS), evidenced by an increased activity of senescence‐associated beta‐galactosidase (SA‐β‐Gal), growth arrest, and overexpression of p21, p16, and p53 mRNAs. Yet, unlike mouse and human fibroblasts, RS and EIS Spalax cells showed undetectable or decreased expression of the well‐known SASP factors: interleukin‐6 (IL6), IL8, IL1α, growth‐related oncogene alpha (GROα), SerpinB2, and intercellular adhesion molecule (ICAM‐1). Apparently, due to the efficient DNA repair in Spalax, senescent cells did not accumulate the DNA damage necessary for SASP activation. Conversely, Spalax can maintain DNA integrity during replicative or moderate genotoxic stress and limit pro‐inflammatory secretion. However, exposure to the conditioned medium of breast cancer cells MDA‐MB‐231 resulted in an increase in DNA damage, activation of the nuclear factor κB (NF‐κB) through nuclear translocation, and expression of inflammatory mediators in RS Spalax cells. Evaluation of SASP in aging Spalax brain and intestine confirmed downregulation of inflammatory‐related genes. These findings suggest a natural mechanism for alleviating the inflammatory response during cellular senescence and aging in Spalax, which can prevent age‐related chronic inflammation supporting healthy aging and longevity.     

The colonic groove or furrow: A comparative morphological study of six species of African mole‐rats (Rodentia,Bathyergidae)

Herbivorous mammals such as nutrias, guinea pigs, chinchillas, and mole‐rats have a longitudinal mucosal colonic groove (furrow) in their ascending colon, which is thought to play a role in the colonic separation mechanism (CSM). It is not known whether this groove is structurally modified to adapt to this function in mole‐rat species. The morphology of this groove was studied in 32 mol‐rats, four species, one of which consisted of three subspecies, endemic to southern Africa and two species found in eastern Africa. The macroscopic morphology of the groove was documented, and samples for histological examination were taken. The groove was wide at its origin at the cecocolic junction and was lined on either side by a row of papillae with the opposing papillae slightly offset in arrangement. The papillated groove gradually decreased in size toward the distal part of the ascending colon where it disappeared. This pattern was similar in all species except in Heterocephalus glaber, where the papillae were absent and the groove was lined by two longitudinal ridges. A histological examination of cross sections revealed that the mucosa covering the inner and outer walls of the groove was rich in mucous‐secreting goblet cells. The walls of the groove contained smooth muscle extending from the inner circular smooth muscle layer at the base to the tips of the papillae in all species examined as well as arteries, lymphatic vessels, and prominent sinusoid‐like veins. The groove could be demonstrated both macroscopically and histologically in three Bathyergus suillus fetuses of varying sizes. The sinusoid‐like veins present in all grooves, regardless of macroscopic shape, suggest that they have a role in the functioning of the groove. J. Morphol. 2009. © 2009 Wiley‐Liss, Inc.     

Superficial,suprahyoid, and infrahyoid neck musculature in naked mole-rats (Heterocephalus glaber): Relative size and potential contributions to independent movement of the lower incisors

Naked mole-rats (Heterocephalus glaber) are fossorial, eusocial rodents that exhibit the unusual capability of moving their lower incisors independently in lateral and rostroventral directions. The evolution of this trait would presumably also involve concurrent alterations in neck musculature to support and control movements of the lower incisors. In order to assess morphological adaptations that might facilitate these movements, we performed detailed dissections of the neck musculature of adult naked mole-rats. In addition to characterizing attachment sites of superficial, suprahyoid, and infrahyoid musculature, we also quantified muscle mass and mandibular features thought to be associated with gape (condyle height, condyle length, and jaw length). Based on muscle attachment sites, the platysma myoides may contribute to lateral movement of the lower incisor and hemi-mandible in naked mole-rats. The large digastric muscle is likely to be a main contributor to rostroventral movement of each lower incisor. The geniohyoid and mylohyoid muscles also likely contribute to rostroventral movements of the lower incisors, and the mylohyoid may also produce lateral spreading of the hemi-mandibles. The transverse mandibular (intermandibularis) muscle likely serves to reposition the lower incisors back to a midline orientation following a movement.     

A cryptic genetic boundary in remnant populations of a long‐lived,bird‐pollinated shrub Banksia sphaerocarpa var. caesia (Proteaceae)

Plant species often exhibit genetic structure at multiple spatial scales. Detection of this structure may depend on the sampling strategy used. We intensively sampled a common, naturally patchy Banksia species within a 200 km² region, in order to assess patterns of genetic diversity and structure at multiple spatial scales. In total, 1321 adult shrubs from 37 geographical populations were genotyped using eight highly polymorphic microsatellite markers developed for the species. Genetic structure was detected at three spatial scales. First, we identified a stark and unexpected division of the landscape into two genetic subregions, one to the north‐east and one to the south‐west of the sampling grid. This differentiation was based on sudden, highly structured differences in common allele frequencies, the cause of which is unknown but that may relate to physical and reproductive barriers to gene flow, localised selection, and/or historical processes. Second, we observed genetic differentiation of populations within these subregions, reflecting previously described patterns of restricted pollen flow in this species. Finally, fine‐scale genetic structure, although weak, was observed within some of the populations (mean S_P = 0.01837). When feasible, intensive sampling may uncover cryptic patterns of genetic structure that would otherwise be overlooked when sampling at broader spatial scales. Further studies using a similar sampling strategy may reveal this type of discontinuity to be a feature of other south‐western Australian taxa and has implications for our understanding of evolution in this landscape as well as conservation into the future. © 2015 The Linnean Society of London, Biological Journal of the Linnean Society, 2015, 115 , 241–255.