Specialized brain regions and sensory inputs that control locomotion in leeches.

Locomotor systems are often controlled by specialized cephalic neurons and undergo modulation by sensory inputs. In many species, dedicated brain regions initiate and maintain behavior and set the duration and frequency of the locomotor episode. In the leech, removing the entire head brain enhances swimming, but the individual roles of its components, the supra- and subesophageal ganglia, in the control of locomotion are unknown. Here we describe the influence of these two structures and that of the tail brain on rhythmic swimming in isolated nerve cord preparations and in nearly intact leeches suspended in an aqueous, "swim-enhancing" environment. We found that, in isolated preparations, swim episode duration and swim burst frequency are greatly increased when the supraesophageal ganglion is removed, but the subesophageal ganglion is intact. The prolonged swim durations observed with the anterior-most ganglion removed were abolished by removal of the tail ganglion. Experiments on the nearly intact leeches show that, in these preparations, the subesophageal ganglion acts to decrease cycle period but, unexpectedly, also decreases swim duration. These results suggest that the supraesophageal ganglion is the primary structure that constrains leech swimming; however, the control of swim duration in the leech is complex, especially in the intact animal.

Developmentally-regulated sodium channel subunits are differentially sensitive to alpha-cyano containing pyrethroids.

Juvenile rats have been reported to be more sensitive to the acute neurotoxic effects of the pyrethroid deltamethrin than adults. While toxicokinetic differences between juveniles and adults are documented, toxicodynamic differences have not been examined. Voltage-gated sodium channels, the primary targets of pyrethroids, are comprised of alpha and beta subunits, each of which have multiple isoforms that are expressed in a developmentally-regulated manner. To begin to test whether toxicodynamic differences could contribute to age-dependent deltamethrin toxicity, deltamethrin effects were examined on sodium currents in Xenopus laevis oocytes injected with different combinations of rat alpha (Na(v)1.2 or Na(v)1.3) and beta (beta(1) or beta(3)) subunits. Deltamethrin induced tail currents in all isoform combinations and increased the percent of modified channels in a concentration-dependent manner. Effects of deltamethrin were dependent on subunit combination; Na(v)1.3-containing channels were modified to a greater extent than were Na(v)1.2-containing channels. In the presence of a beta subunit, deltamethrin effects were significantly greater, an effect most pronounced for Na(v)1.3 channels; Na(v)1.3/beta(3) channels were more sensitive to deltamethrin than Na(v)1.2/beta(1) channels. Na(v)1.3/beta(3) channels are expressed embryonically, while the Na(v)1.2 and beta(1) subunits predominate in adults, supporting the hypothesis for age-dependent toxicodynamic differences. Structure-activity relationships for sensitivity of these subunit combinations were examined for other pyrethroids. Permethrin and tetramethrin did not modify currents mediated by either subunit combination. Cypermethrin, beta-cyfluthrin, esfenvalerate and fenpropathrin all modified sodium channel function; effects were significantly greater on Na(v)1.3/beta(3) than on Na(v)1.2/beta(1) channels. These data demonstrate a greater sensitivity of Na(v)1.3 vs Na(v)1.2 channels to deltamethrin and other cyano-containing pyrethroids, particularly in the presence of a beta subunit.

Activation of two forms of locomotion by a previously identified trigger interneuron for swimming in the medicinal leech.

Higher-order projection interneurons that function in more than one behavior have been identified in a number of preparations. In this study, we document that stimulation of cell Tr1, a previously identified trigger interneuron for swimming in the medicinal leech, can also elicit the motor program for crawling in isolated nerve cords. We also show that motor choice is independent of the firing frequency of Tr1 and amount of spiking activity recorded extracellularly at three locations along the ventral nerve cord prior to Tr1 stimulation. On the other hand, during Tr1 stimulation there is a significant difference in the amount of activity elicited in the ventral nerve cord that correlates with the motor program activated. On average, Tr1 stimulation trials that lead to crawling elicit greater amounts of activity than in trials that lead to swimming.

Modification of leech behavior following foraging for artificial blood.

In this study we examined whether the foraging for artificial blood affected the behavioral responsiveness of leeches to electrical stimulation of the body wall. After foraging for artificial blood, electrical stimulation of the posterior end of the leech significantly increased the percentage of stimulation trials that elicited locomotory activity--swimming and crawling--compared to the behaviors elicited when leeches did not forage or foraged for normal saline. On the other hand, shortening always dominated the behavioral profile of the leech to anterior stimulation even after foraging for artificial blood. In intact anterior end-isolated nerve cord preparations, we also found that application of artificial blood to the intact anterior end was sufficient to modify motor responsiveness to DP nerve stimulation. Full strength artificial blood had an overall negative effect on the likelihood of DP nerve stimulation initiating swimming and on the average length of elicited swim episodes compared to when pond water surrounded the anterior end. Application of a 10% solution of artificial blood to the anterior end led to an increase in the likelihood of DP nerve stimulation eliciting swimming.

An introductory biology lab that uses enzyme histochemistry to teach students about skeletal muscle fiber types.

One important goal of introductory biology laboratory experiences is to engage students directly in all steps in the process of scientific discovery. Even when laboratory experiences are built on principles discussed in the classroom, students often do not adequately apply this background to interpretation of results they obtain in lab. This disconnect has been described at the level of medical education (4), so it should not be surprising that educators have struggled with this same phenomenon at the undergraduate level. We describe a new introductory biology lab that challenges students to make these connections. The lab utilizes enzyme histochemistry and morphological observations to draw conclusions about the composition of functionally different types of muscle fibers present in skeletal muscle. We report that students were not only successful at making these observations on a specific skeletal muscle, the gastrocnemius of the frog Rana pipiens, but that they were able to connect their results to the principles of fiber type differences that exist in skeletal muscles in all vertebrates.