Melvyn D. Goldfinger

Identification of γ-Aminobutyric Acid-Like Immunoreactive Neurons in the Rat Cuneate Nucleus

Neurons in the cuneate nucleus of the rat were examined for gamma-aminobutyric acid-like immunoreactivity (GABA-LI) using antiserum raised against GABA-glutaraldehyde-keyhole limpet hemocyanin. GABA-LI neurons were analyzed for size, shape, and distribution and compared to Nissl-stained neurons. GABA-LI cell bodies were located at all rostral-caudal levels and were distributed randomly throughout the nucleus except at the level of the obex, where they were limited to the peripheral region of the cuneate nucleus. GABA-LI cell bodies had a significantly smaller mean cross-sectional area than the total cuneate neuronal population and comprised 21.5% of the total neuronal population as assessed with Nissl-staining. These results are consistent with the hypothesis that GABA is involved in processing somatosensory information in the rat dorsal column nuclei.

Computation of high safety factor impulse propagation at axonal branch points.

The propagation of single impulses at axonal branch points and widenings was computed numerically. Previous computational studies have held that an action potential propagates across an unmyelinated axonal branch point with diameter-dependent lowered likelihood, such that increasingly complex arborizations could eliminate propagating information. This result is counterintuitive to the principle of information divergence within neuronal circuits. The present study re-examined this result. The boundary conditions at a branch point were extracted from a physical analog circuit with actual branches. The main results were that impulse propagation was reliable past branch points and widenings, and that conduction velocity changed spatially as a function of fiber geometrical inhomogeneity.

HPLC-EC determination of free primary amino acid concentrations in cat cisternal cerebrospinal fluid.

This paper describes an HPLC-EC method for measuring the concentrations of 9 free primary amino acids in cerebrospinal fluid (CSF) withdrawn from the cisterna magna of Nembutal-anesthetized adult cats. Amino acid derivatives were formed with o-phthalaldehyde and beta-mercaptoethanol; subsequently, excess thiol reagent was removed with iodoacetamide. During elution through a C18 5-micron column, the electrochemical detectors sensitivity was switched to accommodate the wide ranges of CSF amino acid concentrations. The analysis was acceptably precise and linear at and above the CSF levels and did not require CSF deproteinization. During the 23 min elution, the concentrations of 8 CSF amino acids were determined: alanine, asparagine, glutamate, glutamine, glycine, serine, taurine, and tyrosine; measurable concentrations were between 1 and 800 microM. The concentration of GABA was below its detection limit (0.5 microM). To assess the ability to detect small concentration increases which might occur due to experimental manipulations, the minimum detectable increments in CSF amino acid concentrations above endogenous levels were determined.

The morphology and distribution of serotonin-like immunoreactive fibers in the cat dorsal column nuclei.

Fibers showing serotonin-like immunoreactivity (5-HT-LI) are demonstrated in the gracile, cuneate and external cuneate nuclei of the cat using avidin-biotinylated horseradish peroxidase and indirect fluorescence immunohistochemical methods. 5-HT-LI fibers are located in all cytoarchitectural subdivisions of the gracile and cuneate nuclei but are restricted to the medial half of the external cuneate nucleus. In all nuclei, 5-HT-LI fibers consist of long, varicose strands showing a wide range in the diameters of both varicosities and intervaricose segments. Results support the concept that serotonergic systems may be involved in the processing of non-noxious somatosensory submodalities.

Detection by HPLC-EC of Primary Amines Recovered in Aqueous Push-Pull Perfusates from Cat Cuneate Nucleus

A method is described for the quantitative detection of primary amines - particularly free amino acids - recovered in aqueous push-pull perfusates obtained from the cat cuneate nucleus. Isoindole derivatives of primary amine groups are formed by precolumn reaction with o-phthalaldehyde and mercaptoethanol. Derivatized sample components are separated and detected using HPLC with electrochemical detection. Of 22 amino acids standards studied individually, 12 were detectable under the conditions described. Variability of elution times and detector output peak heights were less than 2% and less than 10%, respectively. Concentration curves were linear to the 10 picomole order of magnitude. For cuneate nucleus perfusates, samples recovered during continuous peripheral somatosensory stimulation contained detectable derivative levels elevated above those of control samples. Sources of error in data interpretation are discussed.

Superposition of Impulse Activity in a Rapidly Adapting Afferent Unit Model

Mechanosensory afferent units consist of a parent axon, the peripheral axonal arborization, and the branch terminal mechanoreceptors. The present work uses a mathematical model to describe the contribution of a given number of rapidly-adapting mechanoreceptors to the impulse pattern of their parent axon. In the model impulses initiated by any driven mechanoreceptor instantaneously propagate orthodromically and antidromically. The model also incorporates the axonal absolute refractory period as well as ortho-and antidromically elicited recovery cycles. In separate computations, periodic or random (Poisson process) trains of short-duration stimuli at constant amplitude are delivered to a given number (N=2–30) of co-innervated mechanoreceptors. The superposition of component impulse trains always departs from the theoretical ideal (Poisson process). Such departures are attributable to: (i) the number of driven mechanoreceptors, when N is small, (ii) axonal absolute refractory period, during maximal amplitude stimulation, and (iii) antidromic recovery cycles as well as absolute refractoriness, during submaximal-amplitude stimulation. Computations reveal that this “instantaneous reset” model results in the elimination of information extracted by driven mechanoreceptors. Model predictions with Poisson stimulation at varied amplitudes are compared to G-hair afferent unit responses to analogous stimulation. Qualitatively opposite results with respect to parent axonal impulse patterns imply that the axonal arborization is not simply a substrate for impulse propagation from branch terminals to parent axon.

Evidence for Amino Acid Concentration Gradients Between CSF and Extracellular Fluid

A small volume in the extracellular space of the medulla in the anesthetized cat was perfused with cisternal cerebrospinal fluid (CSF) using a push-pull technique. The recovered perfusate was a mixture of pushed CSF and the extracellular fluid. HPLC-EC analysis showed that the concentration of some primary amino acids in recovered perfusate often differed from their concentrations in CSF. These results suggested that amino acid gradients existed between CSF and the extracellular space.

Theoretical studies of impulse propagation in serotonergic axons

Impulse propagation in small-diameter (1–3 μm) axons with inhomogeneous geometry was simulated. The fibres were represented by a series of 3 μm-long compartments. The cable equation was solved for each compartment by a finite-difference approximation (Cooley and Dodge 1966). First-order differential equations governing temporal changes in membrane potential or Hodgkin-Huxley (1952) conductance parameters were solved by numerical integration. It was assumed that varicosity and intervaricosity segments had the same specific cable constants and excitability properties, and differed only in length and diameter. A single long varicosity or a ‘clump’ of 3 μm-long varicosities changed the point-to-point (axial) conduction velocity within as well as to either side of the geometrically inhomogeneous regions. When 2 μm-diameter, 3 μm-long varicosities were distributed over the 1 μm-diameter fiber length as observed in serotonergic axons, mean axial conduction velocity was between that of uniform-diameter 1 and 2 μm fibers, and changed predictably with different cable parameters. Fibers with inexcitable varicosity membranes also supported impulse propagation. These simulations provided a general theoretical basis for the slow (< 1 M/s) conduction velocity attributed to small-diameter unmyelinated varicose axons.

Poisson-Process Electrical Stimulation: Circuit and Axonal Responses

This work describes a simple circuit which generated a highly Poisson-like sequence of pulses. Resistor noise was amplified in three series stages followed by rectification through a relatively large shunt resistance. This yielded a sequence of variable-amplitude transients, which were inverted, amplified with DC adjustment, and fed into a Schmitt trigger/multivibrator chip for pulse generation. The pulse generation frequency was modulated by the amplification of the rectified transients. The stochastic characteristics of the output pulse train were Poisson-like over a wide frequency range, as assessed using the intervent interval distribution and expectation density as steady-state and real-time estimators, respectively. In separate tests, the output pulse train was applied to forelimb cutaneous axons of the anesthetized cat; trains of elicited propagating action potentials were recorded extracellularly from individual G1 axons in the cuneate fasciculus. The stochastic properties of the action potential train differed from those of the stimulus, with longer deadtime, lower mean rate, and an early expectation density peak. These physiological responses to circuit output were similar to those elicited by other generators of Poisson-like stimulation.