Real-time in vivo simultaneous measurements of nitric oxide and oxygen using an amperometric dual microsensor.

This paper reports a real-time study of the codynamical changes in the release of endogenous nitric oxide (NO) and oxygen (O(2)) consumption in a rat neocortex in vivo upon electrical stimulation using an amperometric NO/O(2) dual microsensor. Electrical stimulation induced transient cerebral hypoxia due to the increased metabolic demands that were not met by the blood volume inside the stimulated cortical region. A NO/O(2) dual microsensor was successfully used to monitor the pair of real-time dynamic changes in the tissue NO and O(2) contents. At the onset of electrical stimulation, there was an immediate decrease in the cortical tissue O(2) followed by a subsequent increase in the cortical tissue NO content. The averages of the maximum normalized concentration changes induced by the stimulation were a 0.41 (±0.04)-fold decrease in the O(2) and a 3.6 (±0.9)-fold increase in the NO concentrations when compared with the corresponding normalized basal levels. The peak increase in NO was always preceded by the peak decrease in O(2) in all animals (n = 11). The delay between the maximum decrease in O(2) and the maximum increase in NO varied from 3.1 to 54.8 s. This rather wide variation in the temporal associations was presumably attributed to the sparse distribution of NOS-containing neurons and the individual animal's differences in brain vasculatures, which suggests that a sensor with fine spatial resolution is needed to measure the location-specific real-time NO and O(2) contents. In summary, the developed NO/O(2) dual microsensor is effective for measuring the NO and O(2) contents in vivo. This study provides direct support for the dynamic role of NO in regulating the cerebral hemodynamics, particularly related to the tissue oxygenation.

Cellular basis of neurogenesis in the brain of crayfish, Procambarus clarkii: Neurogenic complex in the olfactory midbrain from hatchlings to adults.

Neurogenesis in the central olfactory pathway of decapod crustaceans persists throughout life. Here we describe the structural basis of neurogenesis within the olfactory deutocerebrum of the crayfish Procambarus clarkii from hatchlings to adults. Using a proliferation marker and immunostaining, we found that throughout development each hemibrain contains a neurogenic complex consisting of five parts: two proliferation zones, each within the neuronal soma clusters containing local or projection interneurons, a tail of proliferating cells extending from each proliferation zone, and an elongated clump of cells where the two tails meet. The clump of cells comprises two subdivisions joined at a nucleus-free central area. Each subdivision consists of a dense group of clump cells with small, spindle-shaped nuclei and is connected to one of the proliferation zones by a strand of fibrous material encompassing the tail of proliferating cells extending from it. We identify one proliferating cell with a large nucleus in each subdivision as a putative neuroblast. Its daughter cells migrate through the strands to the associated proliferation zones, but in the strand leading to the soma cluster of local interneurons this is masked by local proliferation. We conclude that neurogenesis in the olfactory deutocerebrum of juvenile and adult P. clarkii is based on a few neuroblasts that are associated with unique clumps of cells likely representing stem cell niches.

Social domination increases neuronal survival in the brain of juvenile crayfish Procambarus clarkii.

Olfactory cues are among the sensory inputs that crayfish use in establishing dominance hierarchies. Throughout their lives, new neurons are continuously added into brain cell clusters 9 and 10, which contain somata of olfactory local and projection interneurons, respectively. Using markers for DNA synthesis (bromodeoxyuridine) and mitosis (phospho-histone-3), we tested juvenile crayfish (Procambarus clarkii) to examine effects of pairwise social experience on proliferation and survival of cells in these brain regions. Proliferating and mitotic cells appeared within restricted neurogenic areas in both clusters and in ;tails' extending from them. These tails, embedded in tubulin-positive strands, are linked by a patch of cells. Neither cell proliferation nor mitotic activity was affected by social dominance. Cell survival of neuronal precursors was affected by dominance: compared to dominants, subordinates had fewer newborn cells surviving in cluster 9 after 14 days of social experience. Social experience also affected body growth rate, but the effect of social experience on neurogenesis remained when differences in body growth rate were statistically controlled. We conclude that social domination enhances survival of new olfactory interneuronal precursors compared to social subordination but not compared to social isolation.

The effects of social experience on the behavioral response to unexpected touch in crayfish.

Crayfish fight and form a dominance hierarchy characterized by a pattern of repeated agonistic interactions between animals with a consistent outcome of winner and loser. Once a dominance hierarchy is established, dominant animals display an elevated posture with both claws held laterally and forward, whereas subordinate animals display a more prone posture with both claws extended forward and down. Dominant animals behave aggressively towards the subordinate opponent, often approaching and attacking, whereas subordinate animals behave submissively by tailflipping and retreating. To evaluate whether the differences in social behavior are accompanied by differences in responses to non-social stimuli, we exposed socially naïve and experienced crayfish (Procambarus clarkii) to an unexpected touch in different social conditions. Socially naïve animals turned to confront the source of a unilateral touch with raised claws and elevated posture. Dominant animals also turned to face the stimulus source with raised claws and elevated posture, both when tested alone and in the presence of a subordinate opponent. Subordinate animals displayed this orienting response only while separated from their dominant partners. When paired with their dominant partners, subordinates avoided the stimulus source by walking rapidly forwards or backwards. When the subordinate animals were later tested again, first while semi-separated from the dominant and later while fully separated, they displayed a mixed pattern of avoidance and orienting responses. These results indicate that the behavioral responses of subordinate crayfish to touch depend on their social status, their current social conditions and their recent social history.