Development of swimming in the medicinal leech, the gradual acquisition of a behavior.

Observing the development of behavior provides an assay for the developmental state of an embryo's nervous system. We have previously described the development of behaviors that were largely confined to one or a few segments. We now extend the work to a kinematic analysis of the development of swimming, a behavior that requires coordination of the entire body. When leech embryos first begin to swim they make little forward progress, but within several days they swim as effectively as adults. This increase in efficacy depends on changes in body shape and on improved intersegmental coordination of the swim central pattern generator. These kinematic details suggest how the swim central pattern generating circuit is assembled during embryogenesis.

Development of neuronal circuits and behaviors in the medicinal leech.

We are studying the neuronal mechanisms responsible for establishing circuitry underlying the local bending response in the medicinal leech. Local bending replaces an embryonic behavior, circumferential indentation, during the time of initial chemical synaptogenesis in leech embryos. We found that the electrical connections among the motor neurons are established first, about 5% of embryonic time (almost 2 full days) before chemical connections form. The inhibitory connections from muscle inhibitors to muscle excitors are, we hypothesize, responsible for the emergence of local bending. We have also found that the central processes of the excitors--but not the inhibitors--have much longer central processes when their peripheral processes are kept from contacting their target muscles. This system should allow us to test ideas about how individual neurons find their appropriate targets to form functional neuronal circuits.

Staging of middle and late embryonic development in the medicinal leech, Hirudo medicinalis.

We present a description of the last half of embryonic development in the European medicinal leech, Hirudo medicinalis, based entirely on externally visible morphological features, and establish reliably observable stages during that development. Embryogenesis, from the time fertilized eggs are deposited in an eggcase (called a cocoon) to the emergence of juveniles from the cocoon, takes approximately 4 weeks at room temperature. The stages described in this paper extend from the completion of segmentation to the appearance of the final bands of pigmentation. Developmental stages are expressed as percentages of total embryonic developmental time. This staging table was constructed for embryos kept at 20 degrees C. In addition, the development of animals kept at 17 degrees C or at 24 degrees C was compared with those held at 20 degrees C. Development proceeds more quickly at higher temperatures. Because development in embryos held at higher or lower temperatures was linearly related to the stages determined for embryos held at 20 degrees C, the rate of development at any intermediate temperature can be predicted from the staging table at 20 degrees C by simple multiplication.

Development of spontaneous and evoked behaviors in the medicinal leech.

ABSTRACT The ontogeny of behavior in an organism must reflect developmental events in the nervous system, and it thus provides a noninvasive measure of neuronal development. This approach may be particularly fruitful in the medicinal leech because the neuronal basis of several behaviors has been characterized in adult leeches, providing a rich background against which behavioral development can be interpreted. We have investigated the order in which behaviors arise during the period of embryonic development and have determined the time at which each behavior is first expressed. Some behaviors, such as lateral ridge formation, germinal plate bending, spiral twisting, and sidewinding, were produced spontaneously by embryos. Others, such as shortening, circumferential indentation, local bending, and elongation, occurred only when they were elicited by weak mechanical stimulation. Such stimulation rarely evoked a behavioral response in young embryos (at 45% of the time required for complete embryonic development, 45% ED), but by 80% ED embryos responded to nearly 100% of the stimuli presented. In embryos older than 50% ED, the behavior most frequently evoked by stimulation of the anterior end, the posterior end, or the rear sucker was shortening. Stimulation of the midbody usually evoked behavior other than shortening, illustrating that the body was behaviorally compartmentalized, at least in part. Some behaviors observed during embryogenesis are never seen in adult leeches. For example, in response to stimulation of the midbody, young embryos produced a behavior that we have called "circumferential indentation," whereas older embryos produced local bending, a response previously described for adults. The switch from circumferential indentation to local bending may signal the formation of new synaptic connections.

Developmental regulation of segment-specific cholinergic receptors on Retzius neurons in the medicinal leech.

Retzius (Rz) neurons in the midbody ganglia of medicinal leeches responded to ACh, applied to their somata, in a manner that depended upon the neuron's segmental location: Rz neurons in ganglia from midbody segments 5 and 6 [Rz(5,6)] hyperpolarized, whereas Rz neurons from all other segments [Rz(X)] depolarized. Midbody segments 5 and 6 are notable because they contain the male and female reproductive organs. Both types of Rz neurons responded to ACh in a complex way, but the initial phase of each response appeared to be nicotinic because nicotinic agonists evoked the responses and nicotinic antagonists blocked them. The reversal potentials of the responses and the effects of changing the internal and external Cl- concentration indicated that the hyperpolarizing response of Rz(5,6) neurons depended upon Cl- whereas the depolarizing response of Rz(X) neurons did not. The segmentally characteristic responses of Rz neurons arose during embryonic development. Removing the reproductive ducts [the peripheral targets of Rz(5,6)] early in embryogenesis caused the Rz(5,6) neurons to depolarize in response to ACh rather than to hyperpolarize. This result indicates that development of the characteristic response of Rz neurons to ACh is strongly influenced by interactions between the neurons and their appropriate target tissues.

Mesenchyme of embryonic reproductive ducts directs process outgrowth of Retzius neurons in the medicinal leech.

In the two segments of the medicinal leech (Hirudo medicinalis) that contain the male (segment 5) and the female (segment 6) reproductive ducts, the paired Retzius (Rz) neurons are distinguished by several unique properties. For example, the muscles and glands of the body wall are the primary peripheral targets of Rz neurons in standard segments [Rz(X)], whereas the muscles and glands of the reproductive ducts are the primary peripheral targets of Rz neurons in the two reproductive segments [Rz(5,6)]. In this paper, we show that organogenesis and differentiation, which generate an epithelial tube surrounded by mesenchymal cells, occur in the embryonic reproductive ducts at approximately the time when Rz processes first contact these structures. The growth cones leading one branch of the posterior axon of Rz(5,6) contact the duct mesenchymal cells. Following initiation of this contact, these posterior growth cones enlarge and send out numerous filopodia. Secondarily, growth cones leading the anterior axon of each Rz(5,6) also modify their shapes and trajectories. When embryonic reproductive ducts were transplanted into posterior (nonreproductive) segments, the branch of the posterior Rz axon near the ectopic reproductive tissue produced enlarged growth cones and extended several secondary branches into the mesenchyme of the ectopic tissue. This result suggests that the reproductive mesenchyme is attractive to, and can modify the growth of, all Rz neurons. The behavior of Rz(5,6) growth cones suggests that the reproductive mesenchyme cells provide guidance cues that control the location in which Rz axons elaborate their peripheral arborization and form synapses, and that the mesenchyme may also stimulate the production of a densely branched arbor.

Health hazards from oil, soot and metals at a hot forging operation.

An extensive study of a hot forging operation was performed to characterize and quantitate worker exposures to the aerosol formed by an oil-based die lubricant and it's decomposition products. Total particulate breathing zone levels up to 65 mg/m3 and benzo (a) pyrene levels up to 2.9 mug/m3 were measured.