Nathaniel L. Scholz

The NO/cGMP pathway and the development of neural networks in postembryonic lobsters

The nitric oxide/cyclic 3,5-guanosine monophosphate (NO/cGMP) signaling pathway has been implicated in certain forms of developmental and adult neuronal plasticity. Here we use whole-mount immunocytochemistry to identify components of this pathway in the nervous system of postembryonic lobsters as they develop through metamorphosis. We find that the synthetic enzyme for NO (nitric oxide synthase, or NOS) and the receptor for this transmitter (NO-sensitive soluble guanylate cyclase) are broadly distributed in the central nervous system (CNS) at hatching. In the brain, NOS immunoreactivity is intensified during glomerular development in the olfactory and accessory lobes. Whereas only a few neurons express NOS in the CNS, many more neurons synthesize cGMP in the presence of NO. NO-sensitive guanylate cyclase activity is a stable feature of some cells, while in others it is regulated during development. In the stomatogastric nervous system, a subset of neurons become responsive to NO at metamorphosis, a time when larval networks are reorganized into adult motor circuits. cGMP accumulation was occasionally detected in the nucleus of many cells in the CNS, which suggests that cGMP may have a role in transcription. Based on these findings, we conclude that the NO/cGMP signaling pathway may participate in the development of the lobster nervous system. Furthermore, NO may serve as a modulatory neurotransmitter for diverse neurons throughout the CNS.

Expression of nitric oxide synthase and nitric oxide-sensitive guanylate cyclase in the crustacean cardiac ganglion.

The cardiac ganglion is a simple central pattern‐generating network that controls the rhythmic contractions of the crustacean heart. Enzyme assays and Western blots show that whole heart homogenates from the crab Cancer productus contain high levels of nitric oxide synthase (NOS), an enzyme that catalyzes the conversion of arginine to citrulline with concomitant production of the transmitter nitric oxide (NO). Crab heart NOS is calcium‐dependent and has an apparent molecular weight of 110 kDa. In the cardiac ganglion, antibodies to NOS and citrulline indicate the presence of a NOS‐like protein and NOS enzymatic activity in the four small pacemaker neurons and the five large motor neurons of the cardiac network. In addition, all cardiac neurons label positively with an antibody to cyclic guanosine monophosphate (cGMP). The NO donor sodium nitroprusside (SNP, 10 mM) stimulates additional cGMP production in the isolated ganglion. This increase is blocked by [1H](1,2,4)oxadiazole(4,3‐a)quinoxalin‐1‐one (ODQ, 50 μM), an inhibitor of the NO‐sensitive soluble guanylate cyclase (sGC). Taken together, our data indicate that NO‐ and cGMP‐mediated signaling pathways are enriched in the cardiac system relative to other crab tissues and that the cardiac network may be a target for extrinsic and intrinsic neuromodulation via NO produced from the heart musculature and individual cardiac neurons, respectively. The crustacean cardiac ganglion is therefore a promising system for studying cellular and synaptic mechanisms of nitrergic neuromodulation in a simple pattern‐generating network. J. Comp. Neurol. 454:158–167, 2002.

NO/cGMP Signaling and the Flexible Organization of Motor Behavior in Crustaceans

SYNOPSIS. The basic elements of the NO/cGMP signaling pathway have been identified in the nervous systems of animals from nearly all of the major phyla. In crustaceans, the NO/cGMP pathway is associated with certain fundamental neuronal processes, including sensory integration and the organization and production of motor behavior. Here I review the evidence for NO synthesis and action in crustacean neural networks, with an emphasis on the rhythmic motor circuits of the crab stomatogastric ganglion (STG). In the STG, NO appears to be released as an orthograde transmitter from descending projection neurons. NO’s receptor, a cytopasmic isoform of guanylate cyclase (sGC), is expressed in a subset of the cells that participate in the gastric mill and pyloric central pattern generating networks. In spontaneously-active, in vitro preparations of the STG, pharmacological inhibitors of the NO/cGMP pathway cause the two rhythmic motor patterns to collapse into a single conjoint rhythm. Parallel motor output is restored when the ganglion is returned to normal saline. Although precise mechanisms have yet to be determined, these data suggest that NO and cGMP play an important role in the functional organization of STG networks. The STG, as well as other crustacean models, provides a promising context for studying the physiological and behavioral aspects of NO-mediated signaling in the nervous system.

Ecotoxicological Risk of Mixtures

Water pollution is a global environmental challenge that nearly always involves the degradation of aquatic habitats by mixtures of chemical contaminants. Despite this practical reality, environmental regulations and resource management institutions in most countries are inadequate to the task of addressing complex and dynamic combinations of chemicals. Moreover, our scientific understanding of mixture toxicity and the assessment of corresponding risks to aquatic species and communities have not kept pace with worldwide declines in biodiversity or the introduction of thousands of new chemicals into societal use. In this chapter, we review recent research specific to mixtures in three contexts that are broadly applicable to freshwater and marine ecosystems. These include oil spills, urban non-point source pollution, and the agricultural use of modern pesticides. Each of these familiar and geographically extensive forcing pressures is threaded with uncertainty about interactions between contaminants in mixtures. We also briefly consider relevant and often overlapping environmental regulations in the United States and Europe to illustrate why a proactive consideration of chemical mixtures remains elusive in institutional ecological risk assessment. As the case examples show, however, the problem of mixtures is not intractable and targeted research can guide effective conservation and restoration strategies in a chemically complex world.