Mechanisms underlying odorant-induced and spontaneous calcium signals in olfactory receptor neurons of spiny lobsters, Panulirus argus.

We determined if a newly developed antennule slice preparation allows studying chemosensory properties of spiny lobster olfactory receptor neurons under in situ conditions with Ca(2+) imaging. We show that chemical stimuli reach the dendrites of olfactory receptor neurons but not their somata, and that odorant-induced Ca(2+) signals in the somata are sufficiently stable over time to allow stimulation with a substantial number of odorants. Pharmacological manipulations served to elucidate the source of odorant-induced Ca(2+) transients and spontaneous Ca(2+) oscillations in the somata of olfactory receptor neurons. Both Ca(2+) signals are primarily mediated by an influx of extracellular Ca(2+) through voltage-activated Ca(2+) channels that can be blocked by CoCl2 and the L-type Ca(2+) channel blocker verapamil. Intracellular Ca(2+) stores contribute little to odorant-induced Ca(2+) transients and spontaneous Ca(2+) oscillations. The odorant-induced Ca(2+) transients as well as the spontaneous Ca(2+) oscillations depend on action potentials mediated by Na(+) channels that are largely TTX-insensitive but blocked by the local anesthetics tetracaine and lidocaine. Collectively, these results corroborate the conclusion that odorant-induced Ca(2+) transients and spontaneous Ca(2+) oscillations in the somata of olfactory receptor neurons closely reflect action potential activity associated with odorant-induced phasic-tonic responses and spontaneous bursting, respectively. Therefore, both types of Ca(2+) signals represent experimentally accessible proxies of spiking.

Regulation of ephrin-A expression in compressed retinocollicular maps.

Retinotopic maps can undergo compression and expansion in response to changes in target size, but the mechanism underlying this compensatory process has remained a mystery. The discovery of ephrins as molecular mediators of Sperry's chemoaffinity process allows a mechanistic approach to this important issue. In Syrian hamsters, neonatal, partial (PT) ablation of posterior superior colliculus (SC) leads to compression of the retinotopic map, independent of neural activity. Graded, repulsive EphA receptor/ephrin-A ligand interactions direct the formation of the retinocollicular map, but whether ephrins might also be involved in map compression is unknown. To examine whether map compression might be directed by changes in the ephrin expression pattern, we compared ephrin-A2 and ephrin-A5 mRNA expression between normal SC and PT SC using in situ hybridization and quantitative real-time PCR. We found that ephrin-A ligand expression in the compressed maps was low anteriorly and high posteriorly, as in normal animals. Consistent with our hypothesis, the steepness of the ephrin gradient increased in the lesioned colliculi. Interestingly, overall levels of ephrin-A2 and -A5 expression declined immediately after neonatal target damage, perhaps promoting axon outgrowth. These data establish a correlation between changes in ephrin-A gradients and map compression, and suggest that ephrin-A expression gradients may be regulated by target size. This in turn could lead to compression of the retinocollicular map onto the reduced target. These findings have important implications for mechanisms of recovery from traumatic brain injury.

Distribution and function of splash, an achaete-scute homolog in the adult olfactory organ of the Caribbean spiny lobster Panulirus argus.

achaete-scute complex (ASC) genes, which encode basic helix-loop-helix transcription factors, regulate embryonic and adult neurogenesis in many animals. In adult arthropods, including crustaceans, ASC homologs have been identified but rarely functionally characterized. We took advantage of the recently identified crustacean homolog, splash (spiny lobster achaete scute homolog), in the olfactory organ of the Caribbean spiny lobster Panulirus argus to examine its role in adult neurogenesis. We tested the hypothesis that splash is associated with but not restricted to sensory neuron formation in the olfactory organ, the antennular lateral flagellum (LF), of adult spiny lobsters. We demonstrated splash labeling in epithelial cells across LF developmental zones (i.e., proliferation and mature zones), in auxiliary cells surrounding dendrites of olfactory receptor neurons (ORNs), and in immature and mature ORNs, but not in granulocytes or chromatophores. Since ORN proliferation varies with molt stage, we examined splash expression across molt stages and found that molt stage affected splash expression in the ORN mature zone but not in the proliferation zone. In vivo incorporation of bromodeoxyuridine (BrdU) showed no correlation in the cellular pattern of splash expression and BrdU labeling. The intensity of splash labeling was dramatically enhanced in the proliferation zones following LF damage, suggesting enhanced splash expression during repair and/or regeneration. We conclude that splash is not closely associated with the formation of sensory neurons under normal physiological conditions, and we propose that splash is involved in repair and regeneration. We also propose that splash has additional roles other than neurogenesis in adult crustaceans.

Molecular cloning and characterization of homologs of achaete-scute and hairy-enhancer of split in the olfactory organ of the spiny lobster Panulirus argus.

The olfactory organ of the Caribbean spiny lobster Panulirus argus maintains lifelong proliferation and turnover of olfactory receptor neurons (ORNs). Towards examining the molecular basis of this adult neurogenesis, we search for expression of homologs of proneural, neurogenic, and pre-pattern genes in this olfactory organ. We report here a homolog of the proneural Achaete-Scute family, called splash (spiny lobster achaete-scute homolog), and a homolog of the pre-pattern and neurogenic hairy-enhancer of split family, called splhairy (spiny lobster hairy). Semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) indicates a molt stage dependence of the levels of expression of splash and splhairy mRNA in the olfactory organ, with higher expression in premolt than in postmolt or intermolt animals, which is positively correlated with rates of neurogenesis. splash and splhairy mRNA are expressed not only in the olfactory organ but also in other tissues, albeit at lower levels, irrespective of molt stage. We conclude that the expression of achaete-scute and hairy-enhancer of split in the proliferation zone of the olfactory organ of spiny lobsters and their enhanced expression in premolt animals suggest that they play a role in the proliferation of ORNs and that their expression in regions of the olfactory organ populated by mature ORNs and in other tissues suggests that they have additional functions.

Rosette-type tegumental glands associated with aesthetasc sensilla in the olfactory organ of the Caribbean spiny lobster, Panulirus argus.

The lateral antennular flagellum of decapod crustaceans bears unique olfactory sensilla, namely the aesthetascs, and other sensilla types. In this study, we identify a new major tissue in the lateral flagellum of the Caribbean spiny lobster, Panulirus argus, namely "aesthetasc tegumental glands" (ATGs), based on immunostaining with antibodies against CUB serine protease (Csp), in situ hybridization with csp-specific probes, labeling with the F-actin marker phalloidin, labeling with the nuclear marker Hoechst 33258, and staining with methylene blue. Each ATG has 12-20 secretory cells arranged in a rosette. Each secretory cell has a Csp-immunoreactive basal portion and an apical portion containing granular material (metachromatic staining indicative of acid mucopolysaccharides). At the center of each secretory rosette is a phalloidin-positive common locus that gives rise to a main drainage duct projecting toward the cuticle. Scanning electron and light microscopy show that thin ducts traverse the cuticle and connect to "peg pores" proximal to the bases of the aesthetascs, with 3.4 peg pores per aesthetasc. Since the number of common loci is correlated with the number of peg pores, we conclude that each pore represents the outlet of one ATG, and that the secretions are released from them. We conclude further that ATGs and aesthetascs are functionally linked. We hypothesize that ATG secretions have antifouling and/or friction-reducing properties, and that they are spread over the surface of the aesthetascs by antennular grooming. A review of the literature suggests that ATGs are common in decapod crustacean antennules, and that rosette glands and grooming might be functionally coupled in other body areas.

Patterns of neuroAIDS in Africa.

According to UNAIDS, the African population accounts for greater than half of persons infected with HIV. Nevertheless, little information exists characterizing HIV in this population. Thus, the natural history and progression of HIV in the African population is virtually undocumented and therefore, poorly understood. Information regarding virtually every aspect of the disease including microbiology, pathogenicity, virulence, and clinical manifestation is based largely on data from select and limited populations. During the HAART-era, we have seen dramatic and significant changes in patterns of NeuroAIDS in patients in clinical cohorts from the United States and Western Europe. These observations have led to increased understanding of the progression of NeuroAIDS and have improved our ability to design treatment regimens to combat CNS complications resulting from HIV. Despite the existence of antiretroviral therapy for HIV, its absence in Africa along with poor treatments for opportunistic infections associated with HIV have become the main sources of neurological morbidity and mortality. In this context, we are presented with a unique opportunity to cultivate and enhance our understanding of the natural history and progression of NeuroAIDS in the African population thereby, better equipping healthcare providers, patients and their families in addressing this epidemic. This concept is particularly important as rapidly improving and more accessible anti-HIV medications and medications for the treatment of opportunistic infections become available to third world countries such as Africa. We believe that it is imperative to foster research, education and training between institutions in the industrialized world and Africa to close the gap in understanding patterns of NeuroAIDS in Africa.