Neural models of human face perception propose parallel pathways. One pathway (including posterior superior temporal sulcus, pSTS) is responsible for processing changeable aspects of faces such as gaze and expression, and the other pathway (including the fusiform face area, FFA) is responsible for relatively invariant aspects such as identity. However, to be socially meaningful, changes in expression and gaze must be tracked across an individual face. Our aim was to investigate how this is achieved. Using functional magnetic resonance imaging, we found a region in pSTS that responded more to sequences of faces varying in gaze and expression in which the identity was constant compared with sequences in which the identity varied. To determine whether this preferential response to same identity faces was due to the processing of identity in the pSTS or was a result of interactions between pSTS and other regions thought to code face identity, we measured the functional connectivity between face-selective regions. We found increased functional connectivity between the pSTS and FFA when participants viewed same identity faces compared with different identity faces. Together, these results suggest that distinct neural pathways involved in expression and identity interact to process the changeable features of the face in a socially meaningful way.

Although different brain regions are widely considered to be involved in the recognition of facial identity and expression, it remains unclear how these regions process different properties of the visual image. Here, we ask how surface-based reflectance information and edge-based shape cues contribute to the perception and neural representation of facial identity and expression. Contrast-reversal was used to generate images in which normal contrast relationships across the surface of the image were disrupted, but edge information was preserved. In a behavioural experiment, contrast-reversal significantly attenuated judgements of facial identity, but only had a marginal effect on judgements of expression. An fMR-adaptation paradigm was then used to ask how brain regions involved in the processing of identity and expression responded to blocks comprising all normal, all contrast-reversed, or a mixture of normal and contrast-reversed faces. Adaptation in the posterior superior temporal sulcus--a region directly linked with processing facial expression--was relatively unaffected by mixing normal with contrast-reversed faces. In contrast, the response of the fusiform face area--a region linked with processing facial identity--was significantly affected by contrast-reversal. These results offer a new perspective on the reasons underlying the neural segregation of facial identity and expression in which brain regions involved in processing invariant aspects of faces, such as identity, are very sensitive to surface-based cues, whereas regions involved in processing changes in faces, such as expression, are relatively dependent on edge-based cues.

Contralaterally positioned maxillary (upper jaw) venom glands in snakes are mechanically independent, being able to discharge venom from either gland separately. This has led some studies to test venom function and composition of each contralaterally positioned venom gland to investigate any differences. However, the data on the subject to-date derives from limited sample sizes, appearing somewhat contradictory, and thus still remains inconclusive. Here, we tested samples obtained from the left and right venom glands of four N. siamensis specimens for their relative binding to the orthosteric site of amphibian, lizard, snake, bird, and rodent alpha-1 nicotinic acetylcholine receptors. We also show the relative proteomic patterns displayed by reversed phase liquid chromatography - mass spectrometry. Our results indicate that three of the venom gland sets showed no difference in both functional binding and composition, whilst one venom gland set showed a slight difference in functional binding (but not in specificity patterns between prey types) or venom composition. We hypothesise that these differences in functional binding may be due to one gland having previously ejected venom at some time prior to venom extraction, whilst its contralateral counterpart did not. This might cause the differential rate of toxin replenishment to be unequal between glands, thus instigating the difference in potency, likely due to uneven toxin proportions between glands at the time of venom extraction. These results demonstrate that the separate venom producing glands in snakes remain under the same genetic control elements and produce identical venom components.

The binding of compounds to nicotinic acetylcholine receptors is of great interest in biomedical research. However, progress in this area is hampered by the lack of a high-throughput, cost-effective, and taxonomically flexible platform. Current methods are low-throughput, consume large quantities of sample, or are taxonomically limited in which targets can be tested. We describe a novel assay which utilizes a label-free bio-layer interferometry technology, in combination with adapted mimotope peptides, in order to measure ligand binding to the orthosteric site of nicotinic acetylcholine receptor alpha-subunits of diverse organisms. We validated the method by testing the evolutionary patterns of a generalist feeding species (Acanthophis antarcticus), a fish specialist species (Aipysurus laevis), and a snake specialist species (Ophiophagus hannah) for comparative binding to the orthosteric site of fish, amphibian, lizard, snake, bird, marsupial, and rodent alpha-1 nicotinic acetylcholine receptors. Binding patterns corresponded with diet, with the Acanthophis antarcticus not showing bias towards any particular lineage, while Aipysurus laevis showed selectivity for fish, and Ophiophagus hannah a selectivity for snake. To validate the biodiscovery potential of this method, we screened Acanthophis antarcticus and Tropidolaemus wagleri venom for binding to human alpha-1, alpha-2, alpha-3, alpha-4, alpha-5, alpha-6, alpha-7, alpha-9, and alpha-10. While A. antarcticus was broadly potent, T. wagleri showed very strong but selective binding, specifically to the alpha-1 target which would be evolutionarily selected for, as well as the alpha-5 target which is of major interest for drug design and development. Thus, we have shown that our novel method is broadly applicable for studies including evolutionary patterns of venom diversification, predicting potential neurotoxic effects in human envenomed patients, and searches for novel ligands of interest for laboratory tools and in drug design and development.

Glucocorticosteroids (GC) are long-established, widely used agents for induction of remission in inflammatory bowel disease (IBD). Hyperglycaemia is a known complication of GC treatment with implications for morbidity and mortality. Published data on prevalence and risk factors for GC-induced hyperglycaemia in the IBD population are limited. We prospectively characterise this complication in our cohort, employing machine-learning methods to identify key predictors of risk.

Face-selective regions in the amygdala and posterior superior temporal sulcus (pSTS) are strongly implicated in the processing of transient facial signals, such as expression. Here, we measured neural responses in participants while they viewed dynamic changes in facial expression. Our aim was to explore how facial expression is represented in different face-selective regions. Short movies were generated by morphing between faces posing a neutral expression and a prototypical expression of a basic emotion (either anger, disgust, fear, happiness or sadness). These dynamic stimuli were presented in block design in the following four stimulus conditions: (1) same-expression change, same-identity, (2) same-expression change, different-identity, (3) different-expression change, same-identity, and (4) different-expression change, different-identity. So, within a same-expression change condition the movies would show the same change in expression whereas in the different-expression change conditions each movie would have a different change in expression. Facial identity remained constant during each movie but in the different identity conditions the facial identity varied between each movie in a block. The amygdala, but not the posterior STS, demonstrated a greater response to blocks in which each movie morphed from neutral to a different emotion category compared to blocks in which each movie morphed to the same emotion category. Neural adaptation in the amygdala was not affected by changes in facial identity. These results are consistent with a role of the amygdala in category-based representation of facial expressions of emotion.

The ability to recognise familiar faces with ease across different viewing conditions contrasts with the inherent difficulty in the perception of unfamiliar faces across similar image manipulations. Models of face processing suggest that this difference is based on the neural representation for familiar faces being more invariant to changes in the image, than it is for unfamiliar faces. Here, we used an fMR-adaptation paradigm to investigate neural correlates of image-invariant face recognition in face-selective regions of the human brain. Participants viewed faces presented in a blocked design. Each block contained different images of the same identity or different images from different identities. Faces in each block were either familiar or unfamiliar to the participants. First, we defined face-selective regions by comparing the response to faces with the response to scenes and scrambled faces. Next, we asked whether any of these face-selective regions showed image-invariant adaptation to the identity of a face. The core face-selective regions showed image-invariant adaptation to familiar and unfamiliar faces. However, there was no difference in the adaptation to familiar compared to unfamiliar faces. In contrast, image-invariant adaptation for familiar faces, but not for unfamiliar faces, was found in face-selective regions of the medial temporal lobe (MTL). Taken together, our results suggest that the marked differences in the perception of familiar and unfamiliar faces may depend critically on neural processes in the medial temporal lobe.

Prey-selective venoms and toxins have been documented across only a few species of snakes. The lack of research in this area has been due to the absence of suitably flexible testing platforms. In order to test more species for prey specificity of their venom, we used an innovative taxonomically flexible, high-throughput biolayer interferometry approach to ascertain the relative binding of 29 α-neurotoxic venoms from African and Asian elapid representatives (26 Naja spp., Aspidelaps scutatus, Elapsoidea boulengeri, and four locales of Ophiophagus hannah) to the alpha-1 nicotinic acetylcholine receptor orthosteric (active) site for amphibian, lizard, snake, bird, and rodent targets. Our results detected prey-selective, intraspecific, and geographical differences of α-neurotoxic binding. The results also suggest that crude venom that shows prey selectivity is likely driven by the proportions of prey-specific α-neurotoxins with differential selectivity within the crude venom. Our results also suggest that since the α-neurotoxic prey targeting does not always account for the full dietary breadth of a species, other toxin classes with a different pathophysiological function likely play an equally important role in prey immobilisation of the crude venom depending on the prey type envenomated. The use of this innovative and taxonomically flexible diverse assay in functional venom testing can be key in attempting to understanding the evolution and ecology of α-neurotoxic snake venoms, as well as opening up biochemical and pharmacological avenues to explore other venom effects.

Snakes and primates have a multi-layered coevolutionary history as predators, prey, and competitors with each other. Previous work has explored the Snake Detection Theory (SDT), which focuses on the role of snakes as predators of primates and argues that snakes have exerted a selection pressure for the origin of primates' visual systems, a trait that sets primates apart from other mammals. However, primates also attack and kill snakes and so snakes must simultaneously avoid primates. This factor has been recently highlighted in regard to the movement of hominins into new geographic ranges potentially exerting a selection pressure leading to the evolution of spitting in cobras on three independent occasions.

The evolution of an aquatic lifestyle from land dwelling venomous elapids is a radical ecological modification, bringing about many evolutionary changes from morphology to diet. Diet is an important ecological facet which can play a key role in regulating functional traits such as venom composition and prey-specific targeting of venom. In addition to predating upon novel prey (e.g., fish, fish eggs and invertebrates), the venoms of aquatic elapids also face the challenge of increased prey-escape potential in the aquatic environment. Thus, despite the independent radiation into an aquatic niche on four separate occasions, the venoms of aquatic elapids are evolving under convergent selection pressures. Utilising a biolayer interferometry binding assay, this study set out to elucidate whether crude venoms from representative aquatic elapids were target-specific to the orthosteric site of postsynaptic nicotinic acetylcholine receptor mimotopes of fish compared to other terrestrial prey types. Representatives of the four aquatic lineages were: aquatic coral snakes representative was Micrurus surinamensis;, sea kraits representative was Laticauda colubrina; sea snakes representatives were two Aipysurus spp. and eight Hydrophis spp; and water cobras representative was Naja annulata. No prey-specific differences in crude venom binding were observed from any species tested, except for Aipysurus laevis, which showed slight evidence of prey-potency differences. For Hydrophis caerulescens, H. peronii, H. schistosus and M. surinamensis, there was a lack of binding to the orthosteric site of any target lineage. Subsequent testing on the in vitro chick-biventer cervicis muscle preparation suggested that, while the venoms of these species bound postsynaptically, they bound to allosteric sites rather than orthosteric. Allosteric binding is potentially a weaker but faster-acting form of neurotoxicity and we hypothesise that the switch to allosteric binding is likely due to selection pressures related to prey-escape potential. This research has potentially opened up the possibility of a new functional class of toxins which have never been assessed previously while shedding light on the selection pressures shaping venom evolution.

Toxic weaponry in the form of venom and poison has evolved in most groups of animals, including all four major lineages of tetrapods. Moreover, the evolution of such traits has been linked to several key aspects of the biology of toxic animals including life-history and diversification. Despite this, attempts to investigate the macroevolutionary patterns underlying such weaponry are lacking. In this study we analyse patterns of venom and poison evolution across reptiles, amphibians, mammals, and birds using a suite of phylogenetic comparative methods. We find that each major lineage has a characteristic pattern of trait evolution, but mammals and reptiles evolve under a surprisingly similar regime, whilst that of amphibians appears to be particularly distinct and highly contrasting compared to other groups. Our results also suggest that the mechanism of toxin acquisition may be an important distinction in such evolutionary patterns; the evolution of biosynthesis is far less dynamic than that of sequestration of toxins from the diet. Finally, contrary to the situation in amphibians, other tetrapod groups show an association between the evolution of toxic weaponry and higher diversification rates. Taken together, our study provides the first broad-scale analysis of macroevolutionary patterns of venom and poison throughout tetrapods.

Animals that specialise in blood feeding have particular challenges in obtaining their meal, whereby they impair blood hemostasis by promoting anticoagulation and vasodilation in order to facilitate feeding. These convergent selection pressures have been studied in a number of lineages, ranging from fleas to leeches. However, the vampire bat (Desmondus rotundus) is unstudied in regards to potential vasodilatory mechanisms of their feeding secretions (which are a type of venom). This is despite the intense investigations of their anticoagulant properties which have demonstrated that D. rotundus venom contains strong anticoagulant and proteolytic activities which delay the formation of blood clots and interfere with the blood coagulation cascade. In this study, we identified and tested a compound from D. rotundus venom that is similar in size and amino acid sequence to human calcitonin gene-related peptide (CGRP) which has potent vasodilatory properties. We found that the vampire bat-derived form of CGRP (i.e., vCGRP) selectively caused endothelium-independent relaxation of pre-contracted rat small mesenteric arteries. The vasorelaxant efficacy and potency of vCGRP were similar to that of CGRP, in activating CGRP receptors and Kv channels to relax arteriole smooth muscle, which would facilitate blood meal feeding by promoting continual blood flow. Our results provide, for the first time, a detailed investigation into the identification and function of a vasodilatory peptide found in D. rotundus venom, which provides a basis in understanding the convergent pathways and selectivity of hematophagous venoms. These unique peptides also show excellent drug design and development potential, thus highlighting the social and economic value of venomous animals.

Convergence is the phenomenon whereby similar phenotypes evolve independently in different lineages. One example is resistance to toxins in animals. Toxins have evolved many times throughout the tree of life. They disrupt molecular and physiological pathways in target species, thereby incapacitating prey or deterring a predator. In response, molecular resistance has evolved in many species exposed to toxins to counteract their harmful effects. Here, we review current knowledge on the convergence of toxin resistance using examples from a wide range of toxin families. We explore the evolutionary processes and molecular adaptations driving toxin resistance. However, resistance adaptations may carry a fitness cost if they disrupt the normal physiology of the resistant animal. Therefore, there is a trade-off between maintaining a functional molecular target and reducing toxin susceptibility. There are relatively few solutions that satisfy this trade-off. As a result, we see a small set of molecular adaptations appearing repeatedly in diverse animal lineages, a phenomenon that is consistent with models of deterministic evolution. Convergence may also explain what has been called 'autoresistance'. This is often thought to have evolved for self-protection, but we argue instead that it may be a consequence of poisonous animals feeding on toxic prey. Toxin resistance provides a unique and compelling model system for studying the interplay between trophic interactions, selection pressures and the molecular mechanisms underlying evolutionary novelties.

The genus Bitis comprises 17 snake species that inhabit Africa and the Arabian Peninsula. They are responsible for a significant proportion of snakebites in the region. The venoms of the two independent lineages of giant Bitis (B. arietans and again in the common ancestor of the clade consisting of B. gabonica, B. nasicornis, B. parviocula and B. rhinoceros) induce an array of debilitating effects including anticoagulation, hemorrhagic shock and cytotoxicity, whilst the dwarf species B. atropos is known to have strong neurotoxic effects. However, the venom effects of the other species within the genus have not been explored in detail. A series of coagulation assays were implemented to assess the coagulotoxic venom effects of fourteen species within the genus. This study identified procoagulant venom as the ancestral condition, retained only by the basal dwarf species B. worthingtoni, suggesting anticoagulant venom is a derived trait within the Bitis genus and has been secondarily amplified on at least four occasions. A wide range of anticoagulant mechanisms were identified, such as coagulant and destructive activities upon fibrinogen in both giant and dwarf Bitis and the action of inhibiting the prothrombinase complex, which is present in a clade of dwarf Bitis. Antivenom studies revealed that while the procoagulant effects of B. worthingtoni were poorly neutralized, and thus a cause for concern, the differential mechanisms of anticoagulation in other species were all well neutralized. Thus, this study concludes there is a wide range of coagulotoxic mechanisms which have evolved within the Bitis genus and that clinical management strategies are limited for the procoagulant effects of B. worthingtoni, but that anticoagulant effects of other species are readily treated by the South African polyvalent antivenom. These results therefore have direct, real-work implications for the treatment of envenomed patients.

Bites from helodermatid lizards can cause pain, paresthesia, paralysis, and tachycardia, as well as other symptoms consistent with neurotoxicity. Furthermore, in vitro studies have shown that Heloderma horridum venom inhibits ion flux and blocks the electrical stimulation of skeletal muscles. Helodermatids have long been considered the only venomous lizards, but a large body of robust evidence has demonstrated venom to be a basal trait of Anguimorpha. This clade includes varanid lizards, whose bites have been reported to cause anticoagulation, pain, and occasionally paralysis and tachycardia. Despite the evolutionary novelty of these lizard venoms, their neuromuscular targets have yet to be identified, even for the iconic helodermatid lizards. Therefore, to fill this knowledge gap, the venoms of three Heloderma species (H. exasperatum, H. horridum and H. suspectum) and two Varanus species (V. salvadorii and V. varius) were investigated using Gallus gallus chick biventer cervicis nerve-muscle preparations and biolayer interferometry assays for binding to mammalian ion channels. Incubation with Heloderma venoms caused the reduction in nerve-mediated muscle twitches post initial response of avian skeletal muscle tissue preparation assays suggesting voltage-gated sodium (NaV) channel binding. Congruent with the flaccid paralysis inducing blockage of electrical stimulation in the skeletal muscle preparations, the biolayer interferometry tests with Heloderma suspectum venom revealed binding to the S3-S4 loop within voltage-sensing domain IV of the skeletal muscle channel subtype, NaV1.4. Consistent with tachycardia reported in clinical cases, the venom also bound to voltage-sensing domain IV of the cardiac smooth muscle calcium channel, CaV1.2. While Varanus varius venom did not have discernable effects in the avian tissue preparation assay at the concentration tested, in the biointerferometry assay both V. varius and V. salvadorii bound to voltage-sensing domain IV of both NaV1.4 and CaV1.2, similar to H. suspectum venom. The ability of varanid venoms to bind to mammalian ion channels but not to the avian tissue preparation suggests prey-selective actions, as did the differential potency within the Heloderma venoms for avian versus mammalian pathophysiological targets. This study thus presents the detailed characterization of Heloderma venom ion channel neurotoxicity and offers the first evidence of varanid lizard venom neurotoxicity. In addition, the data not only provide information useful to understanding the clinical effects produced by envenomations, but also reveal their utility as physiological probes, and underscore the potential utility of neglected venomous lineages in the drug design and development pipeline.

Understanding drivers of animal biodiversity has been a longstanding aim in evolutionary biology. Insects and fishes represent the largest lineages of invertebrates and vertebrates respectively, and consequently many ideas have been proposed to explain this diversity. Natural enemy interactions are often important in diversification dynamics, and key traits that mediate such interactions may therefore have an important role in explaining organismal diversity. Venom is one such trait which is intricately bound in antagonistic coevolution and has recently been shown to be associated with increased diversification rates in tetrapods. Despite ~ 10% of fish families and ~ 16% of insect families containing venomous species, the role that venom may play in these two superradiations remains unknown.

Snake venom is an adaptive ecological trait that has evolved primarily as a form of prey subjugation. Thus, the selection pressure for toxin diversification is exerted by the prey's physiological targets, with this pressure being particularly acute for specialist feeders, such as the King Cobra species, all of which are snake-prey specialists. However, while extensive research has been undertaken to elucidate key amino acids that guide toxin structure-activity relationships, reciprocal investigations into the specific sites guiding prey-lineage selective effects have been lacking. This has largely been due to the lack of assay systems amenable to systematic amino acid replacements of targeted proteins in the prey's physiological pathways. To fill this knowledge gap, we used a recently described approach based upon mimotope peptides corresponding to the orthosteric site of nicotinic acetylcholine receptor alpha-1 subunits, a major binding site for snake venom neurotoxins that cause flaccid paralysis. We investigated the venoms of four different types of King Cobra (Cambodian, Javan, Malaysian, and Thai). This approach allowed for the determination of the key amino acid positions in King Cobra snake prey that are selectively bound by the toxins, whereby replacing these amino acids in the snake-prey orthosteric site with those from lizards or rats resulted in a significantly lower level of binding by the venoms, while conversely replacing the lizard or rat amino acids with those from the snake at that position increased the binding. By doing such, we identified three negatively charged amino acids in the snake orthosteric site that are strongly bound by the positively charged neurotoxic three-finger toxins found in King Cobra venom. This study, thus, sheds light on the selection pressures exerted by a specialist prey item for the evolution of lineage-selective toxins.