A novel approach to physiology education for biomedical engineering students.

It is challenging for biomedical engineering programs to incorporate an indepth study of the systemic interdependence of cells, tissues, and organs into the rigorous mathematical curriculum that is the cornerstone of engineering education. To be sure, many biomedical engineering programs require their students to enroll in anatomy and physiology courses. Often, however, these courses tend to provide bulk information with only a modicum of live tissue experimentation. In the Electrical, Computer, and Biomedical Engineering Department of the University of Rhode Island, this issue is addressed to some extent by implementing an experiential physiology laboratory that addresses research in electrophysiology and biomechanics. The two-semester project-based course exposes the students to laboratory skills in dissection, instrumentation, and physiological measurements. In a novel approach to laboratory intensive learning, the course meets on six Sundays throughout the semester for an 8-h laboratory period. At the end of the course, students are required to prepare a two-page conference paper and submit the results to the Northeast Bioengineering Conference (NEBC) for consideration. Students then travel to the conference location to present their work. Since the inception of the course in the fall of 2003, we have collectively submitted 22 papers to the NEBC. This article will discuss the nature of the experimentation, the types of experiments performed, the goals of the course, and the metrics used to determine the success of the students and the research.

A push-pull set of elastic strand stretch receptor neurons for the swimmeret in an isopod Bathynomus doederleini.

There coexist two types of neuronal terminal processes attaching to elastic strands at the socket of the swimmeret in Bathynomus doederleini. One of the processes, stretch receptor I is derived from the 1st nerve root of the abdominal ganglion. The other, stretch receptor II is derived from the 2nd nerve root of the ganglion. Both axons of stretch receptors are very thick (30-60 micro m) at sites before the terminal arborization. Cell bodies of the stretch receptors are located in the ganglion of their own segments. The neuronal cell body of the stretch receptor I is located at the anterior half of the hemiganglion ipsilateral to the periphery, and the neuronal cell body of the stretch receptor II at the posterior half of the hemiganglion contralateral to the periphery. Their signaling modalities in response to swimmeret movements were analyzed from intracellular recordings from the cell bodies. Stretch receptor I produced a sustained hyperpolarizing potential in response to protraction of the swimmeret. Stretch receptor II produced a sustained depolarizing potential in response to the protraction, and moreover, generated spike potentials on the rising phase of the depolarizing potential according to its height and steepness. Both the stretch receptors are a push-pull set of elastic strand stretch receptors for the angular position and velocity of swimmeret movements.

Mosaic arrangement of SCP(B-), FMRFamide-, and histamine-like immunoreactive sensory hair cells in the statocyst of the gastropod mollusc Pleurobranchaea japonica.

A pair of statocysts are located in the periganglionic connective tissue of the pedal ganglia of the opisthobranch mollusc Pleurobranchaea japonica. Light- and electron-microscopic observations show that the sensory epithelium of the statocyst consists of 13 disk-shaped hair cells. Each hair cell sends a single axon to the cerebral ganglion through the static nerve. Neurotransmitters in the hair cells were examined by means of immunocytochemistry. Our results show that the 13 sensory hair cells include two SCPB-, three FMRFamide-, and eight histamine-like immunoreactive cells. One hair cell contains a transmitter substance other than SCPB-, FMRFamide, histamine, serotonin, or GABA. One of the two SCPB-like immunoreactive cells, located in the ventral region of the statocyst, is the largest cell in the statocyst. The other, located in the anterodorsal region, shows co-immunoreactivity to both SCPB and FMRFamide antisera. Among the three FMRFamide-like immunoreactive hair cells, one is located in the posteroventral region, separated from the other two, which are adjacent to each other in the anterodorsal region. All the eight histamine-like immunoreactive hair cells are adjacent to one another, occupying the remainder of a triangular pyramid-shaped region. These immunoreactive cells are symmetrically placed in the right and left statocysts. This mosaic arrangement was identical among specimens. Thus the static nerve may code information about position or movement of the statoliths, with the use of different transmitters in the mosaic arrangement of the hair cells.

A reexamination of the synaptic connection between neuron L7 of the abdominal ganglion and neurons of the branchial ganglion in Aplysia californica, A. kurodai and A. juliana.

The presence of a synaptic connection between neuron L7 of the abdominal ganglion and branchial ganglionic neurons (BGNs) was reexamined by means of electrophysiology and fluorescent microscopy using three Aplysia species. We succeeded in recording excitatory postsynaptic potentials produced in a BGN, which followed impulses of L7 one-to-one with a constant latency, even in A. californica as well as in A. kurodai and A. juliana. Dye-injection revealed that fine collaterals extended from a major branch of L7 to the branchial ganglion and arborized in neuropil of the ganglion. The results show that L7 in the three species has a dual function as a motor neuron and as an interneuron for the gill.

Monoamines, amino acids and acetylcholine in the preoptic area and anterior hypothalamus of rats: measurements of tissue extracts and in vivo microdialysates.

A microbore column high-performance liquid chromatography (HPLC) system was used to measure neurotransmitters in tissue extracts and in vivo microdialysates obtained from the preoptic area (PO) and anterior hypothalamus (AH) of rats. The extracts contained norepinephrine, epinephrine, 3,4-dihydroxyphenylacetic acid (DOPAC), dopamine, 5-hydroxyindoleacetic acid (5-HIAA), homovanillic acid (HVA), 5-hydroxytryptamine (5-HT), aspartate, glutamate, GABA, acetylcholine (ACh) and choline. The microdialysates obtained from the PO and AH of freely moving rats contained all of these substances except for norepinephrine, epinephrine, dopamine, and 5-HT. During collection of microdialysate from the PO and AH, core body temperature and locomotor activity were simultaneously measured by means of telemetry. The locomotor activity and body temperature increased during the night. This was accompanied by increased levels of 5-HIAA. The results suggest that serotonergic neuronal mechanisms in the PO and AH may be involved in hypothalamic regulation of spontaneous behaviors and body temperature.

A pharmacological and HPLC analysis of the excitatory transmitter of the cardiac ganglion in the heart of the isopod crustacean Bathynomus doederleini.

In crustacean neurogenic hearts, the myocardium contracts under the tight control of rhythmically active neurons of the cardiac ganglion located inside the heart. We demonstrated that the myocardium of Bathynomus doederleini was sensitive to glutamate, quisqualate, and kainate, and that the tension of myocardial cells developed in a dose-dependent manner. The threshold concentrations were 10(-5) M for quisqualate, 10(-4) M for glutamate, and 3 x 10(-4) M for kainate. Concanavalin A, known to prevent desensitization of glutamate receptors at crustacean neuromuscular junctions, augmented excitatory junctional potentials evoked by the cardiac ganglionic neurons in myocardial cells. Using a high performance liquid chromatography (HPLC), we analyzed glutamate in extracts of the cardiac ganglion and myocardium. We obtained values for glutamate concentrations, 8741.2 +/- 184.2 and 678.2 +/- 10.7 pmol/mg, respectively. Although we attempted to measure monoamines in the extracts by HPLC, these were not detected at measurable (more than 1 fmol per 10-microL sample) levels. In conclusion, myocardial cells in isopod crustaceans were suggested to receive glutamatergic motor innervation.

Evidence for cholinergic inhibitory and serotonergic excitatory neuromuscular transmission in the heart of the bivalve Mercenaria mercenaria

The heart of Mercenaria mercenaria is innervated bilaterally at the atria. A pair of cardiac nerves arise as a branch of the cerebro-visceral connective and run to the posterior end of the junction between each atrium and its efferent branchial vessel. Innervation evidently spreads over the heart, since both inhibitory and excitatory junctional potentials (IJPs and EJPs) can be recorded from the atria, the atrio-ventricular (AV) valve or the ventricle. The cardiac nerves contain inhibitory and excitatory axons. Neural stimulation can cause increases in the frequency or amplitude of beating, depending on the strength and frequency of stimulation. Electrical stimulation of the nerves to the incurrent and excurrent siphons causes bradycardia or tachycardia even after cutting the cerebro-visceral connective at a site anterior to the origin of the cardiac nerves. This may indicate a reflex pathway involving neurons whose cell bodies are located in the visceral ganglion. Neural depression of myocardial action potentials is mediated by discrete IJPs, which follow nerve stimuli one-to-one. IJPs can be recorded from the atria, the AV valve or the ventricle. A long-lasting hyperpolarization follows cessation of excitatory stimulation of the cardiac nerve. IJPs may be produced by cholinergic nerves and are mediated primarily by Cl-. They are blocked by Mytolon and by d-tubocurarine (dTC), but not by methylxylocholine. In low-[Cl-] solution, IJPs are inverted into depolarizing junctional potentials, which are blocked by Mytolon and dTC. Neurally induced tachycardia is mediated by discrete EJPs, which also follow stimuli applied to the cardiac nerve in a one-to-one manner. EJPs can be recorded from the atria, the AV valve and the ventricle. The myocardium and the AV valve were excited by application of serotonin. EJPs recorded from these sites were reduced in amplitude by methysergide (1-methyl-d-lysergic acid butanolamide), suggesting that the EJPs may be serotonergic. Just after entering the heart, at the posterior end of the junction with the efferent branchial vessel, the cardiac nerves contain thick processes which show serotonin-like immunoreactivity. These processes spread along the ramifications of the nerves, which extend to the atrium, the AV valve, the ventricle and even into the wall of the aorta. This study provides direct electrophysiological evidence for serotonergic EJPs and cholinergic IJPs, plus immunocytochemical evidence for neural processes containing serotonin, in the myocardium.

Identification of cardio-inhibitory neurons in the thoracic ganglion of the isopod crustacean Bathynomus doederleini.

We identified cell bodies of a pair of the cardio-inhibitory (CI) neurons in the first thoracic ganglion (TG1) of Bathynomus doederleini. Each of the cell bodies gives rise to a single axon, which runs into the contralateral hemiganglion of TG1 via the commissure and runs to the heart via the fourth root as CI axon. Some fine processes arise from the axon at two points in both hemiganglia.

Identification of cardio-acceleratory neurons in the thoracic ganglion of the isopod crustacean Bathynomus doederleini.

We identified cell bodies of the first and second cardio-acceleratory nerves (CA1 and CA2) in the second and third thoracic ganglia of Bathynomus doederleini. Each cell body gives rise to a single axon, which diverges into two major processes in each case. One of the two major axon processes from the second thoracic ganglion runs to the heart via the third root as CA1, while that from the third thoracic ganglion runs as CA2. The other axon process runs forward to the anterior ganglion in both cases.

Aminergic cellular organization in the gills of Aplysia species.

The constituent elements of the gills of Aplysia kurodai and A. juliana were examined for the presence of biogenic amines using histochemical, immunocytochemical, and HPLC techniques. Aminergic elements were revealed by glyoxylic acid-induced fluorescence in the branchial nerve, branchial ganglion, branchial vessels, and pinnules in both species. Three types of fluorescent cells were found in the neural plexus of the gill in each species. Two of them might be sensory neurons. Although HPLC analysis showed the presence of serotonin and dopamine in all gill structures including fluorescent neural elements, there were regional differences in concentrations of the monoamines. It was noted in the pinnules that there was a much higher concentration of dopamine than serotonin. Serotonin immunocytochemistry revealed neural processes which were immunoreactive to antiserotonin antibody, but serotonin immunoreactivity could not be found in a population of branchioganglionic neuron (BGN) somata. Serotonergic elements in the ganglion may be processes of the central ganglion, while dopaminergic elements may be processes of neurons in the neural plexus, located beyond the branchial ganglion. BGNs were activated by bath-applied dopamine and serotonin. These results suggest that dopaminergic sensory inputs from the neural plexus and serotonergic descending inputs from the abdominal ganglion may be among the inputs received by BGNs. It was found that serotonin depressed excitatory junctional potentials in muscle cells of the efferent branchial vessel, which were induced by an identified neuron of the abdominal ganglion. The aminergic cellular organization of the gill may involve serotonergic presynaptic-inhibitory fibers arising from the abdominal ganglion.