Biomedical subjects

B Oakley

Publications and source records attributed to B Oakley.

At least 37 records · Page 2Linked to original sources

PubMed1993

Lectin identification of olfactory receptor neuron subclasses with segregated central projections.

Our previous studies have demonstrated that the primary olfactory projection in rainbow trout is organized nontopographically; the pattern of termination of olfactory axons in the olfactory bulb is unrelated to the distribution of their cell bodies in the olfactory mucosa. In the present research we have further characterized the organization of this projection by examining the lectin-binding properties of olfactory receptor neurons. The results indicate that in trout, as in mammals, populations of olfactory receptor neurons differ significantly from one another in their carbohydrate "signatures." We have identified subsets of olfactory receptor neurons, specified by unique lectin-binding properties, that are widely distributed and intermingled with the other receptor neurons in the olfactory mucosa and nerve, but that segregate as they enter the olfactory bulb and project to restricted regions of the glomerular layer. This pattern of terminations is bilaterally symmetrical, is remarkably consistent across individuals, and reappears when the primary olfactory projection is reconstituted following transection of the olfactory nerve. As revealed by the carbohydrates on subpopulations of receptor neurons, there is substantial order in the nontopographic projection of olfactory receptor neurons to the olfactory bulb. The functional significance of this organization and the means by which it develops and is maintained remain under investigation.

Animals↗

PubMed1992

Immunocytochemical identification of primary olfactory afferents in rainbow trout.

We have used a combination of techniques to analyze the primary olfactory projection in trout: anterograde tract tracing with horseradish peroxidase (HRP) and immunocytochemistry with antisera to olfactory marker protein (OMP) and to keyhole limpet hemocyanin (KLH). HRP labeling and the OMP antiserum revealed a subset of ciliated receptor neurons with a wide dendrite that lacked the protruding knob found on other receptor neurons. The organization of the primary olfactory axons was clearly revealed by antisera to KLH, which reacted with no other neurons. When visualized with anti-KLH, fascicles of olfactory axons penetrated the basal lamina of the olfactory rosette at scattered sites and converged to form the olfactory nerve. Fascicles within the olfactory nerve traveled parallel to the long axis of the nerve until resorted by extensive intermixing as they entered the olfactory bulb. Within the olfactory bulb, most axons terminated in nine discrete terminal fields in the glomerular layer; however, a few olfactory nerve axons projected into the ventral medial telencephalon. Fascicles supplying each terminal field in the glomerular layer followed distinctive trajectories within the olfactory nerve layer. Axons ending in two terminal fields made brush-like terminations rather than the glomerular terminations characteristic of the remaining seven fields. After unilateral olfactory nerve transection, returning olfactory axons reestablished the normal pattern of terminal fields within 14 weeks. It is likely that the organization of afferents in the trout olfactory bulb is similarly well regulated during normal receptor cell replacement.

Animals↗

PubMed1992

Receptor cell regeneration and connectivity in olfaction and taste.

The capacity of adult mammalian gustatory and olfactory receptor cells to regenerate and make synaptic reconnections provides examples that may be useful in initiating replacement of other kinds of sensory receptor cells. The sensory code for taste quality may not be degraded by taste receptor cell turnover because axons probably recouple to the appropriate type of new receptor cell by axon-receptor cell affinity. Experiments on the development and regeneration of taste receptor cells suggest that they regenerate and turn over by recapitulating the late but not the early steps in taste bud development. To evaluate the replacement of vertebrate olfactory receptors, we began by characterizing the spatial pattern of primary olfactory projections in rainbow trout. Contiguous clusters of HRP-labeled olfactory receptor neurons (ORN) make highly divergent projections to the olfactory bulb. Retrograde transport of fluorescent latex beads revealed that a given restricted site in the glomerular layer received axons from ORNs widely scattered in the epithelium. Hence, ORN axons do not form point-to-point or regional topographic maps. Rather, the olfactory epithelial sheet makes a plane-to-point or holographic-like projection, since any given point in the glomerular layer receives information from the entire olfactory epithelial plane. Receptor cells that reacted with the lectin pokeweed agglutinin were highly dispersed in the olfactory epithelium with axons widely scattered in the olfactory nerve. Yet, as a consequence of the extensive reaggregation of axons at the nerve-bulb interface, the lectin-positive axons fasciculated and converged into a subregion of the glomerular layer.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1992

Spatial buffering of extracellular potassium by Müller (glial) cells in the toad retina.

We examined the role of Müller (glial) cells in buffering light-evoked changes in extracellular K+ concentration, [K+]o, in the isolated retina of the toad, Bufo marinus. We found evidence for two opposing Müller cell current loops that are generated by a light-evoked increase in [K+]o in the inner plexiform layer. These current loops, which are involved in the generation of the M-wave of the electroretinogram (ERG), prevent the accumulation of K+ in the inner plexiform layer by transporting K+ both to vitreous and to distal retina. In addition, under dark-adapted conditions, we found evidence for a Müller cell current loop that is generated by a light-evoked decrease in [K+]o in the receptor layer. This current loop, which is involved in the generation of the slow PIII component of the ERG, helps to buffer the light-evoked decrease in [K+]o throughout distal retina by transporting K+ from vitreous. The spatial buffering fluxes of K+ can be abolished by blocking Müller cell K+ conductance with 200 microM Ba2+. The separate contributions of the M-wave and slow PIII currents to Müller cell spatial buffering were isolated by various pharmacological treatments that were designed to enhance or suppress light-evoked activity in specific retinal neurons. Our results show that Müller cell K+ currents not only buffer light-evoked increases in [K+]o, but also buffer light-evoked decreases in [K+]o, and thereby diminish any deleterious effects upon neuronal function that could arise in response to large changes in [K+]o in the plexiform layers. Moreover, our results emphasize that spatial buffering currents generate many components of the electroretinogram.

Animals↗

PubMed1992

PII component of the toad electroretinogram.

1. The PII component of the electroretinogram (ERG) is comprised of the b-wave and the DC component and is thought to reflect bipolar cell activity. Although the b-wave is generated in large part by a K+/Müller cell mechanism, the origin of the DC component is unclear. In this paper we detail our investigation of the origin of the DC component. We hypothesize that the DC component is generated by a K+/Müller cell mechanism identical to that involved in b-wave generation. 2. We studied the ERG in the dark-adapted, isolated retina preparation of the toad, Bu fo marinus. We used K+ ion-sensitive microelectrodes (K+ISM), as well as conventional intra- and extracellular microelectrodes, to record [K+]o changes, the vitreal ERG, and Müller cell responses. 3. We used the excitatory amino acid receptor agonist N-methyl-DL-aspartate (NMDLA) to inhibit light responses of third-order neurons and thereby eliminate most of the ERG M-wave. In the absence of the M-wave, the ERG consisted of PII and PIII. We then superfused the retina with a solution containing both kynurenic acid (KYN) and 2-amino-4-phosphonobutyric acid (APB), which together inhibit all retinal responses proximal to the photoreceptors. In the presence of KYN and APB, the ERG consisted only of PIII. Using digital subtraction, we reconstructed PII. To our knowledge, this is the first report of the isolation of a PII component in the ERG of a nonmammalian species. 4. Using K+ISMs, we recorded the distal K+ changes in the outer plexiform layer (OPL).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1991

The rate and locus of development of rat vallate taste buds.

Several hundred taste buds develop and mature in the trench walls of the rat's vallate papilla during the first 3 months after birth. The objective of this investigation of taste bud development was to determine: (i) whether the vallate papilla has local growth zones where new taste buds form, (ii) whether new taste buds arise by the division of mature taste buds, (iii) how many days are required for a new taste bud to mature, and (iv) whether a taste pore forms as the taste bud reaches a critical volume. Camera lucida drawings were made of more than 4000 iron hematoxylin-stained, serially sectioned, vallate taste buds. The relative abundance of immature taste buds declined exponentially with age, from 18% of the mature taste buds at day 15 to 2% at day 90. At days 21, 33 and 45 most of the immature taste buds (those lacking a taste pore) were located in growth zones at the anterior and posterior extremes of the vallate trench. A mean of 10.5 days was required for the maturation of each cohort of immature taste buds present at days 15, 21, 33 and 45. Vallate taste buds were added de novo; fission of mature taste buds was rare. Taste buds varied widely in the taste bud volume at which a pore formed and in the final volume of the taste bud.

Animals↗

PubMed1991

Neuronal-epithelial interactions in mammalian gustatory epithelium.

There are two neurally controlled morphogenetic programmes in the fungiform papillae of the rodent tongue. When gustatory axons are present they not only promote the expression of taste receptor cells but also simultaneously suppress the outgrowth of an ectopic filiform spine. Innervation during development is essential for the initial formation of taste buds. Consequently, denervation during development irreversibly prevents the establishment of a permanent line of stem cells for taste buds, in contrast to the reliable re-emergence of taste buds after the reinnervation of denervated adult gustatory epithelium. Monoclonal antibodies to keratins selectively recognize cells in particular differentiated states and may contribute to an understanding of the cell lineages and cell-cell interactions that lead to taste buds and filiform spines. Regeneration may recapitulate the late steps in taste development but not the early steps, which are slower and subject to irreversible alteration with denervation. Arguments are set forth that the nerve dependence of taste buds and some other cutaneous receptor cells provides an important selective advantage. Nerve dependence ensures that a more protective integumental covering will replace any superficial, secondary sensory cells rendered useless by denervation.

Adaptation, Physiological↗

PubMed1991

A deconvolution technique for improved estimation of rapid changes in ion concentration recorded with ion-selective microelectrodes.

In biological preparations, measurements of rapid, stimulus-evoked changes in ion concentration by ion-selective microelectrodes can be distorted by the limited bandwidth of these sensors. Techniques were developed to reconstruct the actual change in ion concentration using deconvolution of the electrode's output signal and the electrode's transfer function. In the vertebrate retina, a knowledge of the actual time course of a light-evoked increase in extracellular K+ concentration was used to provide a rigorous test of a hypothesis regarding the electrical origin of a clinically important component of the electroretinogram.

Animals↗

PubMed1991

M-wave of the toad electroretinogram.

1. In the retina, two distinct, light-evoked releases of K+ have been described. One takes place in the outer plexiform layer (OPL) and is termed the "distal K+ increase." The other takes place in the inner plexiform layer (IPL) and is termed the "proximal K+ increase." Although the distal K+ increase generates the electroretinogram (ERG) b-wave, the contribution of the much larger proximal K+ increase to the ERG is less well understood. In this paper we detail our investigation of the proximal K+ increase and its contribution to the ERG. We describe an ERG component, the M-wave, which had not heretofore been observed in the diffuse-flash, vitreal ERG. 2. We studied the proximal K+ increase and the ERG M-wave in the isolated retina preparation of the toad, Bufo marinus. We used K(+)-sensitive microelectrodes, as well as conventional intra- and extracellular microelectrodes, to record K+ changes, the local (or intraretinal) ERG, the vitreal ERG, and Müller cell responses. 3. As in earlier studies of the amphibian and cat M-wave, we readily observed an M-wave in the intraretinal, or local, ERG (LERG). The M-wave we studied had characteristics similar to those of M-waves that were previously described. Specifically, we found that the M-wave was generated by a Müller cell response to the proximal K+ increase and that both the proximal K+ increase and the LERG M-wave were spatially tuned. 4. We used the aspartate receptor agonist, N-methyl-DL-aspartate (NMA), to reveal that an M-wave is present in the vitreal ERG. Researchers who previously investigated the M-wave were unable to identify an M-wave in the vitreal ERG. We found that the toad ERG M-wave was a small, positive potential that was partially obscured by the much larger b-wave and slow PIII components. 5. We observed that picrotoxin (PTX) had an excitatory effect on inner retina, as evidenced by an enhanced proximal K+ increase and an enhanced M-wave. This result indicates that it is likely that GABAergic inhibition in inner retina plays an important role in retinal processing in the toad. 6. At threshold, we found that the ERG consisted mainly of an M-wave, indicating that the amphibian threshold ERG is driven by proximal retina. This result is analogous to previous observations of the threshold ERG in cat. However, in cat, the M-wave and threshold response have been described as distinct ERG components.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1991

Evaluation of projection patterns in the primary olfactory system of rainbow trout.

Topographic projections are important for coding sensory information in the visual, auditory, and somatosensory systems but are of uncertain importance in the coding of olfactory information. We searched for topographic projections between olfactory receptor cells and the olfactory bulb of the rainbow trout Oncorhynchus mykiss. Anterograde axonal tracing with HRP revealed that the olfactory axons arising from discrete regions of the olfactory epithelium travel together within the olfactory nerve. The abrupt resorting and redistribution of these axons at the interface between the olfactory nerve and olfactory bulb imply that local cues control and organize axonal projections. The sites of termination of HRP-labeled axons in the glomerular layer could not be predicted from the location of their cell bodies in the periphery. Retrograde tracing with fluorescently labeled latex beads, injected into glomerular subregions as small as 1% of the total glomerular volume, labeled receptor cells dispersed throughout the olfactory epithelium. The distributions of labeled receptor cells were uncorrelated with the bulbar injection sites. Double-labeling experiments revealed that even widely separated sites in the glomerular layer receive axons from comingled populations of receptor cells. Hence, the evidence indicates that the spatial arrangement of olfactory receptor cells in the epithelium is not preserved in the termination of their axons in the olfactory bulb. We conclude that the primary olfactory in trout lacks point-to-point or regionally topographic organization and that the entire extent of the olfactory epithelium contributes axons to each region of the glomerular layer.

Animals↗

PubMed1990

Inhibitory interactions among rodent taste axons.

The left side of the tongue of the Mongolian gerbil, Meriones unguiculatus, was experimentally innervated with both chorda tympani nerves. While this dual innervation did not increase the number or volume of fungiform taste buds on the left side, at least half of the taste buds were dually innervated since they could be neurotrophically maintained by either chorda tympani nerve. Impulse discharges occurred simultaneously in the native (left) and foreign (right) chorda tympani nerves when the taste stimulus was restricted to the left side of the tongue. The marked attenuation of the phasic or tonic portions of some taste responses suggested that dual innervation had enhanced inhibition, especially of foreign chorda tympani responses. This was confirmed when electrical stimulation of the native chorda tympani reduced the peak summated action potential discharges of the foreign chorda tympani to NaCl or sucrose by an average of 52 and 41%, respectively. Inhibition began within seconds and continued with an 11.5-min half-life. The inhibition was unaffected by acutely disconnecting either chorda tympani nerve from the brain. We propose that dual chorda tympani innervation accentuated lateral inhibitory connections that may function normally to reduce spurious sensory signals in taste axons.

Animals↗

PubMed1990

Evidence for Na+/H+ exchange in vertebrate rod photoreceptors.

In this paper, we have documented our investigation of pH regulation in the rod photoreceptor of the toad, Bufo marinus. Unlike other neural tissues, the retina depends upon aerobic glycolysis to meet its energy requirements. A consequence of its reliance on glycolysis is a large metabolic production of protons (H+) which must be extruded by pH regulating mechanisms. Based on the work of previous authors, we propose that rods share these same characteristics with whole retina. Our results in rods are consistent with the hypothesis that under nominally bicarbonate-free conditions, an amiloride-sensitive Na+/H+ exchanger contributes to pH regulation in this cell.

Amiloride↗

PubMed1990

Ion-selective microelectrodes suitable for recording rapid changes in extracellular ion concentration.

A method for fabricating double-barrel, ion-selective microelectrodes with fine tips (0.5-1.5 microns) and rapid response times is described. When made into K(+)-selective microelectrodes, the electrodes respond to changes in [K+]o with a time constant of 70-95 ms. The electrical response of these electrodes to common-mode voltages can be made to have a time constant of less than 2 ms, which minimizes electrical artifacts from field potentials. The application of these microelectrodes to the measurement of rapid, transient changes in retinal [K+]o is presented.

Animals↗

PubMed1990

Neural control of ectopic filiform spines in adult tongue.

The tongue surface directly above a fungiform taste bud is flat, thinly keratinized, and free of filiform spines. We examined fungiform papillae in serial sections of rat and gerbil tongues after unilateral transection of the chorda-lingual nerve had caused many fungiform taste buds to degenerate. Such empty fungiform papillae often formed a solitary keratinized outgrowth that closely resembled the spine of an ordinary filiform papilla. By six months an ectopic spine was found on 61% of empty fungiform papillae, but never on fungiform papillae that contained a taste bud. Experimental innervation of the tongue reduced the incidence of ectopic filiform spines in proportion to the cross-sectional area of the trigeminal nerve branches tested (the mylohyoid nerve, the lingual nerve, lingual + mylohyoid or lingual + auriculotemporal nerves). The chorda tympani nerve was 60 times more effective than trigeminal nerves in preventing ectopic filiform spines. We suggest that positive and negative trophic actions are normal characteristics of taste axons, for they promote the formation of taste buds and prevent the expression of ectopic filiform spines. By preventing the outgrowth of ectopic spines on fungiform papillae, taste axons maintain a thinly keratinized apical surface that can be breached by the taste receptor cells.

Animals↗

PubMed1990

K(+)-evoked Müller cell depolarization generates b-wave of electroretinogram in toad retina.

We tested the hypothesis that a light-evoked increase in [K+]o produces a depolarization of the Müller cell membrane, which in turn generates the electroretinogram b-wave current. Using Bufo marinus isolated retinas and K(+)-selective microelectrodes, we recorded two distinct light-evoked increases in extracellular K+ concentration: one in the inner plexiform layer and the other near the outer plexiform layer; the "distal" K+ increase was found over only 10-microns depth and had a maximum amplitude of 0.3 mM. We also recorded the electroretinogram and the light-evoked responses of rods and Müller cells. After correction for the response time of the K(+)-selective microelectrode, the waveforms of all three of these responses were almost exactly as predicted by an earlier computer simulation of the K+/Müller cell hypothesis of the b-wave by Newman and Odette [Newman, E.A. & Odette, L.L. (1984) J. Neurophysiol. 51, 164-182]. The distal K+ increase and the b-wave varied in a similar manner as a function of stimulus irradiance. Superfusion with 0.2 mM Ba2+ attenuated both the Müller cell depolarization and the b-wave by approximately 65% but had no significant effect upon the distal K+ increase. Because Ba2+ reduces K+ conductance of Müller cells, these results are very strong support of the K+/Müller cell hypothesis of the origin of the electroretinogram b-wave; the light-evoked increase in extracellular potassium concentration still is present during superfusion with Ba2+, but the K(+)-evoked Müller cell depolarization and the b-wave are decreased in amplitude because Müller cell K+ conductance is reduced.

Animals↗

PubMed1989

Extracellular pH in the isolated retina of the toad in darkness and during illumination.

1. Extracellular pH (pHo) was measured in the isolated retina preparation of the toad, Bufo marinus, using H(+)-selective microelectrodes. During superfusion with phosphate-buffered solution (pH 7.8), which had a low buffering capacity, pHo in the inner retina was 7.0-7.2 and there was a pHo gradient throughout the distal retina and into the bathing solution. 2. The retinal acidity appears to be due in part to the combined reactions of glycolysis and ATP hydrolysis, since anoxia greatly increased the pHo gradient, while superfusion with either glucose-free pyruvate solution or strophanthidin decreased this gradient. 3. Maintained illumination evoked both an acidification in the proximal retina and an alkalinization in the distal retina. Blocking synaptic transmission to second-order neurones (1.0 mM-aspartate) decreased the acidification but had little effect on the alkalinization, consistent with the notion that the alkalinization is of receptoral origin, while the acidification is of post-receptoral origin. 4. Retinal neurones extrude a significant amount of acid via Na(+)-H+ exchange, since 2.0 mM-amiloride, a blocker of Na(+)-H+ exchange, caused a sustained alkalinization in darkness and decreased the light-evoked changes in pHo, while 1.0 mM-4-acetamido-4'-isothiocyanatostilbene-2.2'-disulphonic acid (SITS), a blocker of Cl(-)-HCO3- exchange, produced a much smaller alkalinization. 5. Switching to a bicarbonate-buffered solution having a 75 times greater buffering capacity than the phosphate-buffered solution caused retinal pHo to become less acidic and significantly decreased the amplitude of the light-evoked pHo changes. 6. Addition of 2.0 mM-acetazolamide, a carbonic anhydrase inhibitor, to the bicarbonate-buffered solution increased both the pHo gradient and the light-evoked changes in pHo. These data are consistent with the idea that carbonic anhydrase, which is concentrated in Müller (glial) cells and to a lesser extent in horizontal cells, increases the effectiveness of the bicarbonate buffer system. 7. Switching from bicarbonate-buffered to phosphate-buffered solutions attenuated the b-wave of the electroretinogram, most likely by acidifying pHo. Overall, our results emphasize the importance of the bicarbonate buffer system in buffering pHo during periods of variable acid extrusion in light and in darkness.

Animals↗

PubMed1987

Neural induction of taste buds.

Bilateral innervation allows more than 80% of the 610 vallate taste buds to survive removal of one IXth nerve in adult rats. Removal of both IXth nerves in neonatal or adult rats results in the absence of taste buds. In studying development, we found that removing or crushing one IXth nerve in three-day-old neonates profoundly decreased the number of vallate taste buds that subsequently developed. Specifically, after removal of one IXth nerve at 3 days, only 228 taste buds formed, compared with 496 taste buds that one nerve would maintain in adults. Thus, during normal development, the right and left IXth nerves interact synergistically, as at least 150 more taste buds develop than predicted by the sum of the independent action of each IXth nerve. This suggests that vallate taste buds are induced by the IXth nerve. A second example of synergism, representing evidence for the neural induction of taste buds, came from experiments in which we crushed the left IXth nerve 3 days after birth and found that these regenerated IXth nerve axons induced 4 times as many taste buds in the presence of the normal right IXth nerve (118 taste buds) as in its early absence (30 taste buds). We conclude that taste buds are neurally induced and that axons of the IXth nerve interact synergistically in inducing them, rather than competing for targets. We propose that in development innervated progenitor cells form stem cells which lead to taste bud cells.

Animals↗

PubMed1987

Postnatal development of the vallate papilla and taste buds in rats.

The postnatal maturation of the vallate papilla and its taste buds was quantitatively investigated in rats by ligh microscopy. Specifically, we measured postnatal increases in the size of mature vallate taste buds and the vallate papilla, increases in the thickness of the gustatory epidermis, and increases in the number of mature taste buds and taste cells per bud. Mature taste buds, defined as those having a taste pore, are rare at birth but proliferate rapidly during the first postnatal month until an average of 610 mature taste buds has accumulated by 90 days. Throughout this postnatal period, mature taste buds adjust to the developmental thickening of the epidermis by continuously increasing in length. Mature taste buds also increase in width, in part due to a threefold increase from 10 and 45 days in the number of taste cells per bud. From 10 to 21 days there is an average daily net increase of three cells per mature taste bud. The maturational increase in taste buds and cells may contribute to the functional changes in taste nerve responses known to occur over the course of several generations of taste receptor cells. The dimensions of the vallate papilla and the surface area of the gustatory epithelium increase logarithmically with age. Although mature taste buds continue to increase in number until 90 days, both taste bud density (178/mm2) and the number of cells per mature taste bud (70-75 cells) reach ceilings by 45 days. Thus, density-dependent factors appear to control vallate taste bud maturation. The immaturity of lingual taste buds in newborn rats supports the view that odor, rather than taste, is the chemosensory signal that guides suckling in altricial rodents.

Animals↗