Dynamical mechanisms of odor processing in olfactory bulb mitral cells

In the olfactory system, the contribution of dynamical properties such as neuronal oscillations and spike synchronization to the representation of odor stimuli is a matter of substantial debate. While relatively simple computational models have sufficed to guide current research in large-scale network dynamics, less attention has been paid to modeling the membrane dynamics in bulbar neurons that may be equally essential to sensory processing. We here present a reduced, conductance-based compartmental model of olfactory bulb mitral cells that exhibits the complex dynamical properties observed in these neurons. Specifically, model neurons exhibit intrinsic subthreshold oscillations with voltage-dependent frequencies that shape the timing of stimulus-evoked action potentials. These oscillations rely on a persistent sodium conductance, an inactivating potassium conductance, and a calcium-dependent potassium conductance and are reset via inhibitory input such as that delivered by periglomerular cell shunt inhibition. Mitral cells fire bursts, or clusters, of spikes when continuously stimulated. Burst properties depend critically on multiple currents, but a progressive deinactivation of I(A) over the course of a burst is an important regulator of burst termination. Each of these complex properties exhibits appropriate dynamics and pharmacology as determined by electrophysiological studies. Additionally, we propose that a second, inconsistently observed form of infrathreshold bistability in mitral cells may derive from the activation of ATP-activated potassium currents responding to hypoxic conditions. We discuss the integration of these cellular properties in the larger context of olfactory bulb network operations.     

The three-dimensional ultrastructure of diabetic glomeruli revealed by the quick-freezing and deep-etching method

The three-dimensional ultrastructure of the glomerular basement membrane (GBM) and mesangial matrix (MM) in streptozotocin (STZ)-induced diabetic rats and in one case of a human diabetic nephropathy were examined using the quick-freezing and deep-etching method. In the diabetic rats, the GBM inner layer was diffusely enlarged, and the meshwork structures in the GBM middle layer and the MM were markedly irregular due to the rupturing of fine fibrils. These irregularities and enlargements of the mesh pores in the diabetic rats developed during the experimental period and were significantly different from those in the control rats. Insulin treatment after STZ injection significantly prevented ultrastructural changes in the GBM and MM. The human diabetic case was a 69-year-old male who had been suffering from non-insulin dependent diabetes mellitus for 29 years. He had a background diabetic retinopathy, microalbuminuria and a decrease in lower limb tendon reflexes. Conventional electron microscopy revealed electron-lucent areas in enlarged subendothelial spaces and mesangium. By the quick-freezing and deep-etching method, the GBM inner layer was diffusely enlarged, and the meshwork structure of the GBM middle layer and the MM showed marked irregularity due to the rupturing of fine fibrils. These findings suggest that the initial morphological changes in diabetic nephropathy include structural abnormalities of fine fibrils in the GBM and MM, which may correspond to the subendothelial and mesangial electron-lucent areas as observed in conventional ultrathin sections.     

Patterns of intracellular potentials in salamander mitral/tufted cells in response to odor stimulation

1. Changes in membrane potential and temporal patterns of spikes were analyzed in 30 output cells in the salamander olfactory bulb in response to stimulation with 1-s pulses of the odorants isoamyl acetate, cineole, and camphor. The odor responses were more complex than responses to electrical stimulation of the olfactory nerve or olfactory tracts, with which they were compared. Most began with hyperpolarization and contained prolonged hyperpolarizing and depolarizing potentials that appeared to be compound postsynaptic potentials. These potentials were related to periods of spike inhibition and excitation. The temporal patterns of the responses resembled S-type (for suppression) and E-type (for excitation) patterns described previously in extracellular-unit studies. 2. In single cells, graded but nonmonotonic changes in the responses were observed with increases in the odor concentration from 10(-3) to 10(-1) vapor-phase saturation. Abrupt changes from one category of temporal response pattern to another were generally not observed in response to different concentrations of a single odorant but were frequently observed when the stimulus was changed from one odorant to another. 3. In S-type patterns, the first event was always membrane hyperpolarization and spike inhibition, regardless of the odor concentration. At all concentrations, simple S-type responses were observed in which a single period of hyperpolarization and inhibition lasted several seconds. At moderate to high concentrations, complex S-type responses were observed in which a period of excitation followed an initial period of hyperpolarization and inhibition. In these responses, spikes were often elicited near the termination of the odor pulse, occasionally as early as 300-400 ms after pulse onset. A prolonged period of inhibition followed the period of excitation. 4. In E-type patterns, the first event depended on the odor concentration. At all concentrations, complex responses were observed in which a period of excitation occurred with short latency, followed by a period of inhibition. At low to moderate concentrations, a brief initial period of hyperpolarization preceded the excitation. This initial period of hyperpolarization was always shorter than those in complex S-type responses to equivalent concentrations. However, the range of spike latencies overlapped that of S-type responses to high concentrations. With increasing odor concentration, spike latencies in the E-type responses decreased relative to the onset and peak of the initial hyperpolarization. At high concentrations. spikes were frequently elicited preceding a single period of hyperpolarization and inhibition.(ABSTRACT TRUNCATED AT 400 WORDS)     

Visuospatial coding in primate prefrontal neurons revealed by oculomotor paradigms

1. Visual responses and their relationship to delay-period activity were studied by recording single neuron activity from the prefrontal cortex of rhesus monkeys while they performed an oculomotor delayed-response (ODR) and a visual probe (VP) task. In the ODR task, the monkey was required to maintain fixation of a central spot of light throughout the cue (0.5 s) and delay (3 s) periods and then make a saccadic eye movement to one of four or eight locations where the visual cue had been presented. In the VP task, the same visual stimuli that were used in the ODR task were presented for 0.5 s, but no response was required. The VP task was thus employed to test the passive visual response and, by comparison with cue-elicited activity in the ODR task, to examine the degree of behavioral enhancement present in prefrontal visual activity. 2. Among 434 neurons recorded from the prefrontal cortex within and surrounding the principal sulcus (PS), 261 had task-related activity during at least one phase of the ODR task, and 74 of these had phasic visual responses to the onset of the visual cues with a median latency of 116 ms. The visual responses of 69 neurons were excitatory, and 5 neurons were inhibited. Five of the neurons with excitatory visual responses also responded transiently after the offset of the cue. 3. Visual responses were classified as directional for 71 PS neurons (96%) in that excitatory or inhibitory responses occurred only for location of cues in a restricted portion of the visual field. Only 3 PS neurons were omnidirectional, i.e., responded equivalently to cues in all locations tested. 4. The best direction and tuning specificity of all PS neurons with directional visual responses were estimated from parameters yielding the best fit to a Gaussian-shaped tuning function. The best direction for the majority (71%) of neurons was toward the visual field contralateral to the hemisphere where the neuron was located. The remaining neurons had their best directions in the ipsilateral field (18%) or along the vertical meridian (11%). 5. The specificity of directional tuning for PS visual responses was quite variable, ranging from neurons that responded only to one of the eight cue locations to neurons that responded to all eight, but in a clearly graded fashion. The standard deviation parameter of the Gaussian curve indexed the breadth of directional tuning of each neuron; its median value was 37 degrees.(ABSTRACT TRUNCATED AT 400 WORDS)     

Coding of odor molecules by mitral/tufted cells in rabbit olfactory bulb. I. Aliphatic compounds.

1. Recordings of extracellular spike responses were made from single mitral/tufted cells in the main olfactory bulb of urethan-anesthetized rabbits. Olfactory epithelium ipsilateral to the recorded olfactory bulb was stimulated with homologous series of aliphatic compounds using periodic artificial inhalations. 2. In the dorsomedial part of the main olfactory bulb, single mitral/tufted cells were activated by subsets of n-fatty acids with similar hydrocarbon chain lengths. Response selectivities of single mitral/tufted cells were examined in detail using a series of n-fatty acids at five different concentrations. The results indicate that although the range of effective fatty acids is broader at the higher concentrations, the best response at higher concentrations was similar to that determined at lower concentrations. 3. Analysis of single-unit responses to the panel of fatty acids, including those with branched hydrocarbon chains, suggested that the determinants for the response specificities of individual mitral/tufted cells in the dorsomedial region include the overall size of hydrocarbon chains of the odor ligand molecules. 4. Single mitral/tufted cells in the dorsomedial region tended to be activated not only by fatty acids but also by n-aliphatic aldehydes. For a panel of a homologous series of n-aldehydes at five different concentrations, individual mitral/tufted cells showed response selectivity to subsets of aldehydes with similar hydrocarbon chain lengths. 5. In most cases, normal aliphatic alcohols and alkanes were ineffective in activating mitral/tufted cells in the dorsomedial region. This suggests that carbonyl group (--C = O) in the odor molecules plays an important role in determining response specificity of these neurons. 6. Examination with an expanded panel of stimulus odor molecules that included ketones and esters indicated that single mitral/tufted cells sensitive to subsets of fatty acids and n-aliphatic aldehydes were also responsive to subsets of ketones and/or esters having hydrocarbon chain lengths similar to those of the effective fatty acids and aldehydes. 7. The present results show a clear correlation between the tuning specificity of individual mitral/tufted cells and the stereochemical structure of the odor molecules, with respect to 1) length and/or structure of hydrocarbon chain, 2) difference in functional group, and 3) position of the functional group within the molecule. 8. A hypothetical diagram suggesting functional convergence of olfactory nerve input to individual glomeruli is proposed to explain the mechanism for selective activation of individual mitral/tufted cells by a range of odor molecules with similar stereochemical structures.     

Disintegration of chromosomes in dead sperm cells as revealed by injection into mouse oocytes

Intracytoplasmic sperm injection of immotile (dead) ejaculated human spermatozoa has been carried out by several centres for the treatment of infertility caused by severe asthenozoospermia (necrozoospermia). No healthy pregnancies have been reported as yet, suggesting irreversible damage to sperm DNA, centrioles and/or other important structures. We investigated this hypothesis by injection of immotile human spermatozoa obtained from several male infertility patients into mouse oocytes and analysis of the oocyte activation rate and sperm chromosome integrity. Motile spermatozoa of the same sample were used as a control. The proportion of living spermatozoa among the immotile was also assessed in each sample and was related to the results of the mouse oocyte injection test. The oocyte activation rate after injection of immotile human spermatozoa into mouse oocytes was the same or only slightly lower than after injection with initially motile spermatozoa (87-100% versus 100% respectively). The rate of normal sperm chromosome spreads correlated significantly (r = 0.90, P < 0.05) with the proportion of living immotile spermatozoa in a given sample. It varied from 4 to 48% for samples containing respectively 8 and 40% of living spermatozoa. Most of the mouse oocytes injected and activated with immotile human spermatozoa were arrested during a prolonged period of time at the interphase of the first cell cycle (from 22 to 60%). Others underwent a delayed nuclear envelope breakdown but showed signs of abnormal structure of both male and female or only the male pronuclear chromosomes. Our data demonstrate an irreversible damage of chromosomes in dead ejaculated human spermatozoa and provide an experimental basis for recommending the use of testicular or epididymal spermatozoa for treatment of male infertility due to necrozoospermia.      

Induction of sister chromatid exchange in preimplantation mouse embryos in vitro by 3H-thymidine or ultraviolet light in combination with caffeine

Preimplantation mouse embryos were exposed in vitro to 3H-thymidine (25, 100, or 250 Bq/ml) or ultraviolet (UV) light (1.35 or 4.05 J/m2), either alone or in combination with caffeine (1 mM with 3H-thymidine and 0.5 mM with UV light). Exposure to 3H-thymidine lasted for 2 days, from the two-cell stage to the late morula/early blastocyst stage, and UV radiation was applied acutely at the late morula/early blastocyst stage. The effects were quantified by the sister chromatid exchange (SCE) assay. All three agents induced SCEs when used singly. 3H-thymidine was effective in inducing SCEs only at 250 Bq/ml, whereas UV light was effective at both fluences. Although caffeine did not induce SCEs when it was added before exposure to bromodeoxyuridine (BrdUrd), which is used to visualize SCEs, it did induce SCEs when present during the entire culture period (3H-thymidine experiments) or during incubation in BrdUrd (UV experiments). Caffeine markedly enhanced the SCE-inducing effect of UV light but did not influence the effect of 3H-thymidine.     

Proctolin-related material in the mouse brain as revealed by immunohistochemistry

By use of a specific antiserum against the insect peptide proctolin we were able to identify proctolin-immunoreactive neurons in the mouse brain. These nerve cells belong to the nuc. mesencephalicus n. trigemini. Furthermore, the antiserum stained very few nerve fibers with varicosities in the immediate neighborhood of the roof of the third ventricle. The chemical identity of the immunoreactive material with genuine proctolin remains to be elucidated.     

The postnatal development of the junctional complexes of the mouse Sertoli cells as revealed by freeze-fracture

Mouse testes of newborn to adult were examined by freezefracture. Between the newborn Sertoli cells, gap junctions consisting of aggregations of the intramembranous particles (about 8 nm in diameter) are frequently found. Some of the junctions are about 1 μm in diameter and show particle-free regions in the aggregation. Linear arrangements of a few particles, which appear to be the initial formation of the occluding junctions, are seen in the newborn Sertoli cells. The occluding junctions are arranged in a meshwork, in which the gap junctions are situated between the stages of newborn to six days of age. The particles of the occluding junctions are predominantly located on the B face in the center of the groove instead of the A face of the ridge. The occluding junctions do not appear to surround the entire circumference of the Sertoli cell of the 6-day-old mouse. The gap junctions decrease in size. In later stages, many parallel occluding junctions (up to forty in number) are found over one Sertoli cell surface and are distributed circumferentially around the entire cell surface, indicating establishment of the blood-testis barrier. The occluding junctions dominate and the gap junctions diminish in number as development proceeds.     

The kinetics of in vivo sister chromatid exchange induction in mouse bone marrow cells by alkylating agents: cyclophosphamide

Administration of cyclophosphamide (5, 10, 20 and 25 mg/kg body weight) to male CD-1 mice 2 hr after subcutaneous implantation of a 5-bromo-2'-deoxyuridine (BrdUrd) pellet (55 mg) resulted in a dose-dependent increase in sister chromatid exchanges (SCE) in bone marrow cells. Treatment with cyclophosphamide (15 mg/kg body weight) at the time of BrdUrd implantation and 2, 6.5, and 13 hr post-BrdUrd implantation resulted in the induction of approximately 19 SCE/cell indicating that the bone marrow SCE response was independent of the time of administration. Treatment with cyclophosphamide (15 mg/kg body weight) at 26, 19, 13, and 6 hr prior to BrdUrd implantation resulted in baseline SCE (3.3 SCE/cell) at 26 hr with an increasing number of SCE/cell with decreasing time prior to BrdUrd implantation. These results compare favorably with those obtained by Kram et al [1981] with mitomycin C (MMC) using a similar protocol. The time-dependent induction of SCE is qualitatively similar for CP and MMC, both of which are bifunctional alkylating agents metabolically activated by oxidation and reduction, respectively, and suggests that these two compounds may induce SCE by a similar mechanism.     

Ultrastructural heterogeneity of "minimal changes" in the kidney glomeruli, detected by light optics

The ultrastructural study of "minimal changes" found at the light microscopic examination of renal biopsies from patients with nephrotic syndrome revealed heterogeneity of morphological changes. A number of diseases included into this group manifest clinically as nephrotic syndrome: hypoplastic dysplasia, lipoid nephrosis, focal-segmentary glomerular hyalinosis, membranous, mesangiomembranous, mesangioproliferative glomerulonephritis. Each of these diseases has its own pathogenetic mechanism resulting in a specific morphological manifestation and consequently in a specific treatment.     

In vivo persistence of sister chromatid exchanges (SCE) induced by gamma rays in mouse bone marrow cells

The sister chromatid exchange (SCE) frequencies induced in bone marrow cells by in vivo irradiation with gamma rays before or after bromodeoxyuridine (BrdUrd) incorporation were compared. The frequency of SCE at different postirradiation times was also measured in bone marrow cells in vivo, irradiated before BrdUrd incorporation. Increased sensitivity to SCE induction by radiation was found in cells after BrdUrd incorporation for one cycle when compared with cells irradiated before BrdUrd incorporation. The increased SCE frequency persisted for at least 72 hr after the initial irradiation, implying that the gamma ray-induced lesion(s) capable of eliciting an SCE are persistent and cannot be easily repaired.     

Bergmann glial development in the mouse cerebellum as revealed by tenascin expression

Tenascin, an astroglia-derived extracellular matrix molecule, is also expressed by radial glia of the embryonic mouse cerebellum. Expression of tenascin can thus be applied as a marker of astroglial development from an early stage, especially prior to the expression of the glial fibrillary acidic protein (GFAP) that can be detected in the postnatal cerebellum. The development of Bergmann glia, specialized cerebello-cortical astroglia with radial processes, was examined by tenascin immunohistochemistry and non-radioactive in situ hybridization histochemistry for tenascin mRNA in the developing mouse cerebellum. Tenascin-immunopositive radial glial processes extending from the ventricular zone to the pia mater retracted toward the cortex in the embryonic cerebellum and occupied a position corresponding to the Bergmann glial processes at the perinatal stage. Tenascin gene-expressing cells were generated in the ventricular zone of the cerbellar primordium and migrated radially toward the cortex. They were stratified in the layer of Bergmann glial somata at the early postnatal stage. They extended GFAP-immunopositive radial processes from the somata to the pia mater as revealed by double-labeling employing tenascin in situ hybridization histochemistry and GFAP-immunostaining. Bergmann glia are therefore considered to develop from cerebellar radial glia by migration of their somata and retraction of their processes. The tenascin gene-expressing cells displayed mitotic activity after alignment in the cortex as revealed by double-labeling by tenascin in situ hybridization histochemistry and immunohistochemical detection of the incorporated bromodeoxyuridine. The above findings suggest that the Bergmann glia in the cortex represent one of the origins of the astroglia in the developing cerebellum.     

Synthesis of lambda light chain subtypes by stimulated and unstimulated mouse B cells

Inbred mouse make 3 lambda chain subtypes. The lambda 1 and lambda 3 chains have similar variable (V) regions (in both the same V gene segment [V lambda 1] is used), whereas lambda 2 and lambda 3 have similar constant (C) regions. Despite the lambda 1 and lambda 3 V region similarity, lambda 1 occurs much more frequently than lambda 3 (and lambda 2) in the serum immunoglobulins and antibody responses of most inbred strains of mice. To explore the basis for the lambda 1 predominance, we compared the rates of synthesis of the 3 subtypes and the frequencies of the B cells that synthesize them, focussing on "resting" (i.e., unstimulated) and on polyclonally stimulated B cells from spleens of unimmunized BALB/c mice. In resting cells the relative rates of synthesis and the relative frequencies of the respective B cells were in accord, indicating that the rate of lambda chain synthesis is approximately the same per resting B cell, regardless of the lambda subtype it produces. However, in the polyclonally stimulated cells, lambda 1 was made 7 times faster than lambda 2 and 10 times faster than lambda 3; normalizing these rates by the frequencies of the respective stimulated cells suggests that in stimulated B cells lambda 1 chains are made 5 times faster per cell than lambda 2 or lambda 3, while the latter are made at about the same rate per cell. In view of the marked structural homology between lambda 2 and lambda 3 genes in segments other than the V-gene segment, we suggest that the pronounced differences among polyclonally stimulated B cells in expression of the genes for the various lambda subtypes may be due to the presence of less potent enhancer-like sequences in the lambda 2 and lambda 3 genes than in the lambda 1 gene.     

Trout immunoglobulin populations differing in light chains revealed by monoclonal antibodies

Monoclonal antibodies (MAbs) raised against trout immunoglobulin (Ig) recognize two independent Ig populations differing in the molecular weight of L chains. The trout Ig population recognized by MAb 2H9 represented congruent to 20% of the total Ig and contained L chains congruent to 26 kDa, whereas the trout Ig population defined by MAb 2A1 comprised 11% of the total Ig and possessed L chains congruent to 24 kDa. The determinants recognized by these MAbs reside on L chains as determined by ELISA using mildly reduced H and L chains. The size heterogeneity of the L chains was not due to differences in glycosylation. Partial peptide maps showed structural differences between both L chains. Both Ig populations were found in each of the 30 trout sera tested. These data extend the existence of L chains diversity in fish, recently described in catfish, to other teleosts.