Discussion

In the present study, we report, for the first time, detailed aspects of swimming behavior of the Nematostella vectensis planula larvae across systematically chosen developmental timepoints and examine its relationship to concomitantly appearing cellular and molecular features. Larval behavior is changing over time, leading to optimized dispersal through the integration of external and internal sensory information that enables the animal to utilize their form in a more efficient way. While some aspects remain relatively stable over development, such as minimum and maximum speed, length of motile cilia, and their beating frequency, the larvae instead shift on the scale of possibilities toward more linear trajectories and faster speeds through the polarization and sharpening of sensory functions. The advance of the sensory systems allows animals to be more active and “sensitive” in response to external stimuli such as light and mechanical stimuli. Additionally, a reafference system based on neuronally mediated body shape contractions is optimized for higher speed and more linear swimming. This sensory integration makes Nematostella larvae versatile swimmers with a rich repertoire of behavioral modes.

For an animal with the size of the Nematostella larvae, swimming in aqueous medium is energetically extremely costly. Indeed, ciliated epithelial cells are metabolically highly active,33 and the ciliary length might be perfectly adapted to swimming efficiency in small organisms.45 Given that a helical swimming pattern is stereotyped across several marine species, some aspects of swimming are potentially also confined by physical laws that determine the maximal efficiency of movement and optimize flow fields.15,27,46,47 While Nematostella swimming speed (0-3 mm/s in still water) and its body size may not endow the animal with the capacity to resist larger currents, their swimming ability is well adapted to facilitate dispersal and habitat selection within their natural ecological niche of tidal pools and marshes, where water may remain still or experience incremental increases. Responding to sensory stimuli by increasing both activity and fine-tuning behavior, reflected in the ability to control swimming modes, facilitates dispersal and enables the larvae to invest their efforts with maximal efficiency. Higher activity in larvae has been observed after mechanical stimulation or in response to short wavelengths of light.11,30 In contrast, responses to stimuli can also induce the opposite response, including drastic body contraction and the cessation of various types of activity.6,13 Constraining dispersal time both during development and also acutely, as observed for Nematostella planula larvae, is an adaptive behavior for an anemone that lives in a relatively restricted area,48 where large dispersal distances could lead to a loss of surviving offspring to the vast ocean currents.

Furthermore, sensory perception will also include self-sensing in the form of proprioception and flow sensing, leading to optimization in swimming gait.37 The ability to actively and effectively utilize the set of swimming modes available to the animals, and to transition between those modes at appropriate timepoints, seems to be governed by an intrinsic mechanism that “turns on” at a certain developmental stage. This increase in sensory abilities co-occurs with increasing numbers of TRP channels in epithelial cells and concomitant elongation of the apical tuft. Being able to access the external and internal sensory information allows the animal to be activated through appropriate stimulation and to optimize swimming gait to reach their goals in the ontogenetically available time.

How cnidarian larvae sense their environment is not well understood. Recent investigations into the cellular and molecular architecture of different cnidarian planulae have, however, opened the possibility for detailed investigations of such aspects.20,21,39,49 In agreement with previous observations that have shown that the apical organ is not required for the larvae-polyp transition21,22 and is lost in some animals prior to settlement,18 our data suggest a sensory role of the apical tuft mainly in swimming behavior. Larvae display larger activation through mechanical input and more linear trajectories at ages that show a prominent tuft, while, at competency, when most animals begin the search for a suitable habitat, the tuft is degrading or lost. Several studies have found sensory receptors expressed in the apical organ19,32 and, together with reports documenting the apical tuft to move as a unit during swimming with a sweeping movement,17,18 it is likely to possess a tactile-sensory or stirring purpose, providing clear polarization of the elongated planula larvae that creates a sensory directionality to guide swimming movement. The increase in tuft length with age might further increase sensory perception by allowing more receptors to be housed in an individual cilium, which would increase the sensitivity of this organ.50 Which other sensory modalities Nematostella larvae use and how the ciliated cells communicate among each other and with the nervous system will be of great interest in future investigations.

In addition to the tuft cilia, our analysis reveals that larvae might not just lose the tuft, but possibly a larger fraction of sensory-motile cilia that enable swimming behavior in the planula larvae. These epithelial cells show expression of functional ion channels only during the motile stage of the larvae, such as Nav2.4 and TRP channels, strongly suggesting that these cells might serve a sensory-motor purpose too. Indeed, previous reports have found sensory-motor cells in cnidarian planula larvae,33 and it is well known that most cilia can sense external stimuli. How and if these ciliated cells and their cognate molecular machinery are involved in the larvae’s swimming is unclear at this time.

A second aspect of improved swimming control is the development of the nervous system. When and how the first nervous systems appeared, and what their role was, is under much active debate. It has been suggested that the nervous system in ciliated metazoan larvae first appeared for signal amplification and direct sensory motor connections.51 We find that many sensory receptors and other “neuronal” ion channels such as voltage-gated sodium channels are expressed in the ciliated epithelium, possibly in some form of larval-specific sensory-motor unit. This ciliated sensory-motor unit appears to be in large parts independent of the nervous system, since blocking synaptic transmission only partially impairs swimming and mostly affects the fine-tuning of behaviors that are mediated by muscular contractions. In other cnidarian larvae, it has been observed that the nervous system is absent or rudimentary and that ciliary sensory motor units might be the only means of steering the animal,33 a feature that is maintained from more basal ciliated larvae.51

We suggest here that the function of the nervous system is the optimized orientation of the sensory organ and possibly other ciliary structures, such as the swimming cilia, which may be controlled by muscular contractions and relaxations. Such active reorientation of the ciliated epithelium might be used to position ciliary rows along the proper body axes, producing more efficient thrust irrespective of CBF. Previous studies have shown that body shape and ciliation can be modified for dispersal efficiency52 and that the generation of coherently directed flow requires cilia to be optimally oriented.53 The body shape and the resulting sensory experience influence the swimming ability and connect appropriate speeds with specialized behaviors. Shape-shifting has been reported in several cnidarian larvae,6,9,54 and further analysis might shed more light on how exactly the complex body shape patterns observed here serve specialized functions in the larva.

Limitations to the study

In the present study, we systematically characterized the swimming behavior of the planula larvae as the animals develop, to describe some key aspects of their behavior. Future studies should venture into cellular and molecular aspects of physiology and sensory perception as well as other types of behaviors. Our study contains some caveats, specifically with respect to the subcellular physiology of cnidarians, which is not well described. How and if different subsystems (muscle, cilia, and neurons) exactly work in concert to optimize signal perception and processing to produce optimal locomotion is likely complex and cannot fully be solved with a generic channel blocker such as MgCl2. Targeted manipulation will require in-depth study of cellular physiology and molecular pharmacology.

Furthermore, the behavioral assays used in our study contain caveats and limitations. In the microscopy behavior, for example, it is difficult to determine exact speed and axis ratio when larvae turn in the vertical plane, and it would of course be optimal to film the animal in all dimensions to avoid data loss. Irrespective of these limitations, our study provides important insights into sensory and motor behavior in cnidarian larvae and provides a basis on which to build more detailed knowledge with targeted manipulations.

Comparative studies can advance our knowledge on how the planula larvae disperse and subsequently find a permanent settlement site with respect to their ecosystem and its physical characteristics. Understanding such details of this critical phase in the larva’s life is important to solve questions related to the current climate crisis and loss of biodiversity. To protect ecosystems, we must understand them. Nematostella vectensis has served as a pioneering model in the biology of cnidarians, and it is urgently required that we develop more tools to understand the fundamental aspects of cnidarian biology.