Teaching About Sex and Gender in Neuroscience: More Than Meets the "XY".

Offering courses on the neuroscience of sex and gender can help support an inclusive curriculum in neuroscience. At the same time, developing and teaching such courses can be daunting to even the most enthusiastic educators, given the subject's complexities, nuances, and the difficult conversations that it invites. The authors of this article have all developed and taught such courses from different perspectives. Our aim is to provide educators with an overview of important conceptual topics as well as a comprehensive, but non-exhaustive, guide to resources for teaching about sex/gender in neuroscience based on our collective experience teaching courses on the topic. After defining vital terminology and briefly reviewing the biology of sex and sex determination, we describe some common topics within the field and contrast our current nuanced understandings from outdated misconceptions in the field. We review how (mis)representation of the neuroscience of sex/gender serves as a case study for how scientific results are communicated and disseminated. We consider how contextualization of sex/gender neuroscience research within a broader historical and societal framework can give students a wider perspective on the enterprise of science. Finally, we conclude with a brief discussion on how to choose learning goals for your course and implementation notes.

Everything in Modulation: Neuromodulators as Keys to Understanding Communication Dynamics.

Across the animal kingdom, the ability to produce communication signals appropriate to social encounters is essential, but how these behaviors are selected and adjusted in a context-dependent manner are poorly understood. This question can be addressed on many levels, including sensory processing by peripheral organs and the central nervous system, sensorimotor integration in decision-making brain regions, and motor circuit activation and modulation. Because neuromodulator systems act at each of these levels, they are a useful lens through which to explore the mechanisms underlying complex patterns of communication. It has been clear for decades that understanding the logic of input-output decision making by the nervous system requires far more than simply identifying the connections linking sensory organs to motor circuits; this is due in part to the fact that neuromodulators can promote distinct and temporally dynamic responses to similar signals. We focus on the vocal circuit dynamics of Xenopus frogs, and describe complementary examples from diverse vertebrate communication systems. While much remains to be discovered about how neuromodulators direct flexibility in communication behaviors, these examples illustrate that several neuromodulators can act upon the same circuit at multiple levels of control, and that the functional consequence of neuromodulation can depend on species-specific factors as well as dynamic organismal characteristics like internal state.

A novel degree of sex difference in laryngeal physiology of Xenopus muelleri: behavioral and evolutionary implications.

Characterizing sex and species differences in muscle physiology can contribute to a better understanding of proximate mechanisms underlying behavioral evolution. In Xenopus, the laryngeal muscle's ability to contract rapidly and its electromyogram potentiation allows males to produce calls that are more rapid and intensity-modulated than female calls. Prior comparative studies have shown that some species lacking typical male features of vocalizations sometimes show reduced sex differences in underlying laryngeal physiology. To further understand the evolution of sexually differentiated laryngeal muscle physiology and its role in generating behavior, we investigated sex differences in the laryngeal physiology of X. muelleri, a species in which male and female calls are similar in rapidity but different with respect to intensity modulation. We delivered ethologically relevant stimulus patterns to ex vivo X. muelleri larynges to investigate their ability to produce various call patterns, and we also delivered stimuli over a broader range of intervals to assess sex differences in muscle tension and electromyogram potentiation. We found a small but statistically significant sex difference in laryngeal electromyogram potentiation that varied depending on the number of stimuli. We also found a small interaction between sex and stimulus interval on muscle tension over an ethologically relevant range of stimulus intervals; male larynges were able to produce similar tensions to female larynges at slightly smaller (11-12 ms) inter-stimulus intervals. These findings are consistent with behavioral observations and present a previously undescribed intermediate sex difference in Xenopus laryngeal muscle physiology.

Generation, Coordination, and Evolution of Neural Circuits for Vocal Communication.

In many species, vocal communication is essential for coordinating social behaviors including courtship, mating, parenting, rivalry, and alarm signaling. Effective communication requires accurate production, detection, and classification of signals, as well as selection of socially appropriate responses. Understanding how signals are generated and how acoustic signals are perceived is key to understanding the neurobiology of social behaviors. Here we review our long-standing research program focused on Xenopus, a frog genus which has provided valuable insights into the mechanisms and evolution of vertebrate social behaviors. In Xenopus laevis, vocal signals differ between the sexes, through development, and across the genus, reflecting evolutionary divergence in sensory and motor circuits that can be interrogated mechanistically. Using two ex vivo preparations, the isolated brain and vocal organ, we have identified essential components of the vocal production system: the sexually differentiated larynx at the periphery, and the hindbrain vocal central pattern generator (CPG) centrally, that produce sex- and species-characteristic sound pulse frequencies and temporal patterns, respectively. Within the hindbrain, we have described how intrinsic membrane properties of neurons in the vocal CPG generate species-specific vocal patterns, how vocal nuclei are connected to generate vocal patterns, as well as the roles of neurotransmitters and neuromodulators in activating the circuit. For sensorimotor integration, we identified a key forebrain node that links auditory and vocal production circuits to match socially appropriate vocal responses to acoustic features of male and female calls. The availability of a well supported phylogeny as well as reference genomes from several species now support analysis of the genetic architecture and the evolutionary divergence of neural circuits for vocal communication. Xenopus thus provides a vertebrate model in which to study vocal communication at many levels, from physiology, to behavior, and from development to evolution. As one of the most comprehensively studied phylogenetic groups within vertebrate vocal communication systems, Xenopus provides insights that can inform social communication across phyla.

Probing forebrain to hindbrain circuit functions in Xenopus.

The vertebrate hindbrain includes neural circuits that govern essential functions including breathing, blood pressure and heart rate. Hindbrain circuits also participate in generating rhythmic motor patterns for vocalization. In most tetrapods, sound production is powered by expiration and the circuitry underlying vocalization and respiration must be linked. Perception and arousal are also linked; acoustic features of social communication sounds-for example, a baby's cry-can drive autonomic responses. The close links between autonomic functions that are essential for life and vocal expression have been a major in vivo experimental challenge. Xenopus provides an opportunity to address this challenge using an ex vivo preparation: an isolated brain that generates vocal and breathing patterns. The isolated brain allows identification and manipulation of hindbrain vocal circuits as well as their activation by forebrain circuits that receive sensory input, initiate motor patterns and control arousal. Advances in imaging technologies, coupled to the production of Xenopus lines expressing genetically encoded calcium sensors, provide powerful tools for imaging neuronal patterns in the entire fictively behaving brain, a goal of the BRAIN Initiative. Comparisons of neural circuit activity across species (comparative neuromics) with distinctive vocal patterns can identify conserved features, and thereby reveal essential functional components.

Evolution of Courtship Songs in Xenopus : Vocal Pattern Generation and Sound Production.

The extant species of African clawed frogs (Xenopus and Silurana) provide an opportunity to link the evolution of vocal characters to changes in the responsible cellular and molecular mechanisms. In this review, we integrate several robust lines of research: evolutionary trajectories of Xenopus vocalizations, cellular and circuit-level mechanisms of vocalization in selected Xenopus model species, and Xenopus evolutionary history and speciation mechanisms. Integrating recent findings allows us to generate and test specific hypotheses about the evolution of Xenopus vocal circuits. We propose that reduced vocal sex differences in some Xenopus species result from species-specific losses of sexually differentiated neural and neuromuscular features. Modification of sex-hormone-regulated developmental mechanisms is a strong candidate mechanism for reduced vocal sex differences.

Species-specific loss of sexual dimorphism in vocal effectors accompanies vocal simplification in African clawed frogs (Xenopus).

Phylogenetic studies can reveal patterns of evolutionary change, including the gain or loss of elaborate courtship traits in males. Male African clawed frogs generally produce complex and rapid courtship vocalizations, whereas female calls are simple and slow. In a few species, however, male vocalizations are also simple and slow, suggesting loss of male-typical traits. Here, we explore features of the male vocal organ that could contribute to loss in two species with simple, slow male calls. In Xenopus boumbaensis, laryngeal morphology is more robust in males than in females. Larynges are larger, have a more complex cartilaginous morphology and contain more muscle fibers. Laryngeal muscle fibers are exclusively fast-twitch in males but are both fast- and slow-twitch in females. The laryngeal electromyogram, a measure of neuromuscular synaptic strength, shows greater potentiation in males than in females. Male-specific physiological features are shared with X. laevis, as well as with a species of the sister clade, Silurana tropicalis, and thus are likely ancestral. In X. borealis, certain aspects of laryngeal morphology and physiology are sexually monomorphic rather than dimorphic. In both sexes, laryngeal muscle fibers are of mixed-twitch type, which limits the production of muscle contractions at rapid intervals. Muscle activity potentiation and discrete tension transients resemble female rather than male X. boumbaensis. The de-masculinization of these laryngeal features suggests an alteration in sensitivity to the gonadal hormones that are known to control the sexual differentiation of the larynx in other Xenopus and Silurana species.

Distinct neural and neuromuscular strategies underlie independent evolution of simplified advertisement calls.

Independent or convergent evolution can underlie phenotypic similarity of derived behavioural characters. Determining the underlying neural and neuromuscular mechanisms sheds light on how these characters arose. One example of evolutionarily derived characters is a temporally simple advertisement call of male African clawed frogs (Xenopus) that arose at least twice independently from a more complex ancestral pattern. How did simplification occur in the vocal circuit? To distinguish shared from divergent mechanisms, we examined activity from the calling brain and vocal organ (larynx) in two species that independently evolved simplified calls. We find that each species uses distinct neural and neuromuscular strategies to produce the simplified calls. Isolated Xenopus borealis brains produce fictive vocal patterns that match temporal patterns of actual male calls; the larynx converts nerve activity faithfully into muscle contractions and single clicks. In contrast, fictive patterns from isolated Xenopus boumbaensis brains are short bursts of nerve activity; the isolated larynx requires stimulus bursts to produce a single click of sound. Thus, unlike X. borealis, the output of the X. boumbaensis hindbrain vocal pattern generator is an ancestral burst-type pattern, transformed by the larynx into single clicks. Temporally simple advertisement calls in genetically distant species of Xenopus have thus arisen independently via reconfigurations of central and peripheral vocal neuroeffectors.