Social insects show a variety of temperature-guided behaviors. Depending on whether heat reaches the sensillum via air movements (convective heat) or as radiant heat, specific adaptations of thermo-sensitive sensilla are expected. In the present study the morphology and the physiology of thermo-sensitive peg-in-pit sensilla (S. coeloconica) of the leaf-cutting ant Atta vollenweideri were investigated. S. coeloconica are located predominantly in a single cluster on the apical antennomere, and connect to the outside through a small aperture. The sensory peg is double-walled, embedded in a chamber and innervated by three unbranched dendrites. Using tungsten electrodes, activity of the sensory neurons was measured. In most cases, the neuron with the largest spike amplitude responds to changes in air temperature (convective heat) as well as to radiant heat. In response to a drop in air temperature, the neuron shows a phasic-tonic response followed by a complete adaptation within 1 min (cold-sensitive neuron). Based on their morphology and physiology, it is suggested that the S. coeloconica are involved in the recently described thermal orientation behavior of A. vollenweideri leaf-cutting ants.

Various microscopic techniques allow investigating structures from submicron to millimeter range, however, this is only possible if the structures of interest are not covered by pigmented cuticle. Here, we present a protocol that combines clearing of pigmented cuticle while preserving both, hard and soft tissues. The resulting transparent cuticle allows confocal laser-scanning microscopy (CLSM), which yields high-resolution images of e.g. the brain, glands, muscles and fine cuticular structures. Using a fluorescent dye, even single labeled neurons can be visualized and resolved up to an imaging depth of 150 μm through the cleared cuticle. Hydrogen-peroxide, which was used to clear the cuticle, does not preclude immunocytochemical techniques, shown by successful labeling of serotonin-immunoreactive neurons (5HT-ir) in the ants' brain. The 'transparent insect protocol' presented here is especially suited for small arthropods where dissection of organs is very demanding and difficult to achieve. Furthermore, the insect organs are preserved in situ thus allowing a more precise three-dimensional reconstruction of the structures of interest compared to, e.g., dissected or sectioned tissue.

The reproductive cycles, growth and survival of the 2 most abundant color morphs (blue and purple) of the colonial ascidian Cystodytes occurring in the western Mediterranean were studied over a period of 20 mo. Previous reports have assigned the 2 morphotypes to the nominal species C. dellechiajei. A seasonal pattern emerged, in which larval release occurred in late spring and summer, followed by a period of active growth. Significant differences between morphotypes were also detected: the cycles of both reproduction and growth rates of the purple morph lagged 1 to 2 mo behind those of the blue morph. Within morphotypes, the time course of reproductive activity and growth rates also displayed a temporal lag that suggests partitioning of resources to either reproduction or growth. In particular, the growth rates of the 2 morphotypes peaked when reproductive activity was at its lowest. There were no significant differences in mortality between these forms, and in both cases a significant negative correlation was found between mortality and size. In the blue morph, maximal growth rate was negatively correlated with size, whilst no significant relationship between growth rate and size was detected in the purple morph. There were no significant differences in growth between colonies of the purple morph that were in contact with the toxic sponge Crambe crambe and those that were not. The differences found in the reproductive cycles of these 2 morphotypes match previously reported genetic and chemical divergences.

The antennae of leaf-cutting ants are equipped with sensilla coeloconica that house three receptor neurons, one of which is thermosensitive. Using convective heat (air at different temperatures), we investigated the physiological characteristics of the thermosensitive neuron associated with the sensilla coeloconica in the leaf-cutting ant Atta vollenweideri. The thermosensitive neuron very quickly responds to a drop in temperature with a brief phasic increase (50 ms) in spike rate and thus classifies as cold receptor (ambient temperature = 24°C). The short latency and the brief phasic response enable the thermosensitive neuron to follow temperature transients up to an estimated frequency of around 5 Hz. Although the neuron responds as a cold receptor, it is extremely sensitive to warm stimuli. A temperature increase of only 0.005°C already leads to a pronounced decrease in the resting activity of the thermosensitive neuron. Through sensory adaptation, the sensitivity to temperature transients is maintained over a wide range of ambient temperatures (18-30°C). We conclude that the thermosensitive neuron of the sensilla coeloconica is adapted to detect minute temperature transients, providing the ants with thermal information of their microenvironment, which they may use for orientation.

We explored the ability of leaf-cutting ants (Atta vollenweideri) to learn the location of a food reward by using thermal information as an orientation cue. During training of single workers, the conditioned stimulus was a distant thermal source placed frontally, 15 mm away from a platform having a leaf fragment as reward. After training, single workers were confronted with the choice between two sides, one being coupled, in a pseudo-randomized design, with a thermal stimulus heated 5 degrees C above environmental temperature. After 10 learning trials, workers significantly chose the side with the thermal stimulus. This showed that workers can use thermal information for spatial orientation in the context of foraging, which may help them to locate, for instance, highly attractive sun-exposed leaves. Thermal radiation alone as orientation cue was sufficient to allow learning, since preclusion of thermal convection during training and test did not impair workers' response. Shielding of both thorax and gaster from the thermal source did not weaken learning, suggesting the sole participation of head and antennae in temperature reception. A thermal stimulus heated 1 degrees C above environmental temperature could not be used as a learned orientation cue, even when foragers were allowed to directly contact the thermal source.