In association with a previously established shift in ecology and behavior across life stages in net-casting spiders - in which mature males no longer engage in an unusual foraging strategy (i.e. net-casting) - we find associated changes in peripheral sensory system structure as well as relative lower and higher-order central nervous system processing investment. Mature male D. spinosa possess decreased posterior median eyes (PMEs) and enlarged anterior median eyes (AMEs) compared to penultimate males, penultimate females, and mature females. Central investment matches peripheral alterations; mature males invest relatively less in lower-order optic neuropils (ONPs) and the mushroom bodies (MBs) - a higher-order integration center that receives direct input from the PMEs, but not the AMEs32,33,34. When compared to penultimate and mature females, mature males also possess relatively larger arcuate bodies (AB) – a higher-order integration center receiving direct input from the AMEs, and likely indirect input from the PMEs32,33,34. Lastly, we describe a species-wide, positive relationship between relative investment in ONPs and the MBs, while also uncovering a negative relationship, and potential trade-off, in relative investment between the ONPs and the AB.

We first confirm a decrease in PME diameter among mature male D. spinosa, when compared to their penultimate stadium, reinforcing previous research showing similar reductions in an Australian net-casting spider, D. subrufa26. Similar to D. subrufa, the absolute size of mature male PMEs in D. spinosa decreases by ~25%. The PMEs in D. spinosa have recently been shown to be important in nocturnal foraging28. As mature males no longer engage in net-casting, they no longer rely on these enlarged eyes for this foraging function and coincident with this, we observe a decrease in PME diameter. Blest and Land26 found that decreased PME diameters relate to a decrease in retinal size, as mature male D. subrufa possess a retina of smaller area, comprised of smaller photoreceptors. While we would predict retinal area to match PME diameter in D. spinosa, and that photoreceptor size varies across groups, these aims were unfortunately beyond the scope of this study. These predictions remain to be studied in D. spinosa.

Consequent to a decrease in PME diameter, we found that mature males possess AMEs with increased diameter when compared to all other focal groups. The AMEs are image-forming in most web-spinning and ground-dwelling spiders, though typically of low quality resolution27, while jumping spiders possess extraordinary AMEs capable of forming highly resolved images30,31. However, nothing is currently known about the potential function of the enlarged AMEs in mature male D. spinosa. While the role of the principal eyes has remained unstudied in Deinopis, the discovery of enlargement of AMEs in mature males suggests a potentially important function of vision for these spiders; and specifically, the type of vision provided by the AMEs. Though mature males no longer rely on PME vision to forage, they may benefit from AME vision while searching for mates. Given that males actively wander in search of mates upon sexual maturation (pers. obs.), enlarged AMEs may aid mature males in locating and/or courting females through enhanced detail discrimination. Another possibility is that AMEs aid males in predator detection. The leg morphology and diurnal posture of mature male D. spinosa is less cryptic than that of mature females and/or juveniles (pers. obs.), potentially increasing selection on effective predator detection. These hypotheses remain to be tested.

In conjunction with differences in peripheral sensory morphology, specifically the reduction in PME diameter, we found a corresponding decrease of relative investment in lower-order visual processing in mature males. When compared to all other focal groups mature males had the lowest relative investment in ONPs, while all other spiders had similar degrees of investment. Neurons are among the most metabolically expensive bodily tissues; as neural tissue requires nearly a magnitude more energy per unit weight to function than most other tissue38,39,40. Additionally, apart from the energetic costs associated with neural function, metabolic costs are also high regarding neural tissue’s maintenance4, leading to an expectation of decrements in brain regions of decreased importance. This is exactly what we observe in D. spinosa, as a decrease in PME diameter is associated with a decreased relative investment of ONPs. Similar within-individual changes in neural investment have been observed in other animals. Seasonal changes in the brains of song birds, for example, are a well-documented case neural plasticity in which brain regions associated with call production become greatly reduced in the winter months when song production ceases41,42,43,44. For mature male D. spinosa, decreasing investment in PME size and processing could benefit males by saving energy to increase longevity or devote to locomotion, thus increasing the chances of finding a mate; and/or by freeing up resources to invest in other, newly important bodily tissue and/or brain regions.

In addition to our observed differences in ONP volume for mature males, relative volume of both of our focal higher-order integration brain regions (MBs and AB) also differed by sex, life stage, or both. As the MBs only receive direct input from the secondary eyes34, we expected PME size to predict relative investment in the MBs, and our analyses matched our predictions. Mature males had the smallest relative MBs, and both penultimate and mature females had greater relative investment in MBs than penultimate males, results that mirror PME size. As females possess larger PMEs than age-matched males, increased relative investment in the MBs might purely be dictated by processing from PMEs. While spider MBs are termed “higher-order integration regions”32, in contrast to insect mushroom bodies35, the functional relevance of spider MBs on behavior is not well-understood32. As such, it is difficult to further interpret our observed differences in relative MB size between male and female D. spinosa.

Similar to the MBs, we also documented differences in relative AB investment across D. spinosa. Relative investment in the AB was greatest in mature males and significantly greater in mature males than in penultimate and mature females spiders. While the AB likely receives information from the secondary eyes indirectly34, it is the only higher-order integration center to receive direct input from the AMEs32,33,34. Thus, similar to the MBs, increased relative investment in the AB could be partially explained by an increase in respective peripheral sensory structures (i.e. AMEs). As mentioned previously, we do not currently know what the function(s) might be of the larger AMEs of mature males. However, beyond AME visual information, the AB also receives inputs from midbrain neurons, potentially leading to a multi-modal integration role32. Mate-searching in spiders is thought to be predominantly chemically-based45,46,47,48. If the AB is, in part, responsible for processing and integrating chemical cues, increased importance of chemical processing in mate-searching males could explain AB investment differences in D. spinosa. To investigate such a hypothesis, it will be important for future studies to quantify the details of chemical processing pathways in net-casting spiders, as well as to explore male reliance on chemical cues for mate-searching. Additionally, it will be important to keep in mind that the AB is also presumed to be a motor control center32, and might function in this capacity as mature males are actively searching for mates. In any sense, future investigations focusing on the AB’s possible role in mate-searching have great potential for yielding interesting and valuable neuroethological results.

In addition to the previously discussed observed differences across life stage and sex with respect to specific processing centers (i.e. the ONPs, MBs, and AB), we also see relationships among these centers that suggest potential synergies, as well as investment trade-offs. Specifically, when comparing relative ONP size with relative MB and AB size, we unveiled contrasting relationships. Relative ONP volume was positively correlated with relative MB volume, while relative ONPs volume was negatively correlated with relative AB volume. Though MBs and the AB are higher-order integration centers, they differ in their connections with visual inputs (among others)33,34. In Cupiennius spiders, the MBs are a third-order optic neuropil that integrates visual information from the secondary eyes; they do not receive direct inputs from the AMEs33,34. If lower and higher-order processing of vision from the same eyes are positively related, then the positive relationship between relative investment in the ONPs and the MBs points to a high degree of processing of secondary eye vision in the ONPs. As the PMEs are the largest eyes of D. spinosa, the bulk of the ONPs are likely associated with their processing. Thus, a positive relationship between the ONPs and the MBs might be expected. In contrast, the AB in Cupiennius receives direct inputs from the AMEs33. Thus, if the ONPs are mostly dedicated to PME processing, this potential trade-off between relative investment in ONPs and the AB might infer a trade-off between distinct types of visual processing (principal vs. secondary).

Following Healy and Rowe49, the current study makes use of intra-specific comparisons, across similarly sized individuals, regarding relative investment in sensory processing. Such comparisons forego multiple difficulties common to comparative neuroanatomy when comparing brain volume across individuals. However, the current study can be regarded as a first step towards better understanding the neuroethology of spiders, and net-casting spiders, specifically. Volumetric analysis is a valuable method in informing researchers where to focus more detailed investigations. As such, our study created various new hypotheses concerning both behavior and neuroanatomy, illustrating the utility of such research. As well, our study displays μCT scanning as a newly-emerging methodology ideal to supplement classical neurohistological methods50. This relatively new method of scanning arthropod internal anatomy has great potential for yielding fast, accurate results across many understudied taxa, opening the door to much greater understanding of how neural systems have evolved.

In summary, we have documented plasticity in both peripheral sensory structures and associated processing centers in the net-casting spider Deinopis spinosa. This work provides an important foundation for more directed hypothesis-testing concerning functional benefits of distinct sensory structures (e.g. AMEs), as well as brain regions (MBs and the AB). Though many questions remain unanswered – e.g. what is the function of the enlarged AMEs of mature male D. spinosa? – we have provided a solid groundwork upon which to build. The neuroethology of spiders remains largely unexplored, especially as compared to the plethora of neuroethological studies conducted on their arthropod relatives – the insects18,51,52,53,54,55– making foundational studies such as this incredibly valuable. We are encouraged by recent strides in spider brain research56,57,58,59.

Numerous studies have shed light on how sensory systems can evolve across animals contrasting in habitat and behavior. Interspecific comparisons have been important in producing hypotheses regarding sensory system evolution, while investigations using intraspecific comparisons of sensory systems across life-stage can shed light on the important, yet understudied, function of sensory system plasticity. Plasticity in both peripheral sensory structures and their underlying neural tissues likely benefit animals exhibiting extraordinary behavioral changes across development, allowing for increased efficiency regarding tissue investment, but how constrained is this developmental plasticity? Future studies such as this, that leverage unusual natural histories and non-model organisms, will contribute to our working knowledge of the costs, benefits, and potential constraints of developmental sensory system plasticity.