Serum steroid hormones and the reproductive cycle of the female bonnethead shark, Sphyrna tiburo

The bonnethead shark Sphyrna tiburo reproduces by placental viviparity with one of the shortest gestation periods (4.5-5 months) known in sharks. In southwest Florida, mating in this species occurs in November, sperm is stored until ovulation/fertilization the following March-April, and parturition occurs in August. Serum concentrations of four steroid hormones (17 beta-estradiol, progesterone, testosterone, and dihydrotestosterone) were determined by radioimmunoassay over a complete reproductive cycle in mature females from a wild population. Serum 17 beta-estradiol and testosterone levels are high during mating and preovulatory stages. Preovulatory concentrations of testosterone are greater in female S. tiburo than in any other female elasmobranch previously studied. Progesterone levels are significantly elevated during preovulatory, ovulatory, and postovulatory stages, while serum dihydrotestosterone levels increase significantly during the preovulatory stage. Our study is the first to demonstrate a sustained rise in progesterone during gestation in a placental shark and suggests a regulatory role for this hormone during the period prior to implantation of the embryos in the uterus.     

The importance of calmodulin in the accessory olfactory bulb in the formation of an olfactory memory in mice

Female mice form an olfactory memory to the pheromones of the mating male, during a critical period after mating. Failure to form this memory results in the male being treated as strange, and hence, his pheromones block pregnancy. Previous studies have shown that formation of this memory is dependent on synaptic mechanisms in the accessory olfactory bulb. A number of studies have pointed to calmodulin as a critical mediator of synaptic plasticity. In this study we have examined the effects of local infusions of drugs which block calmodulin-regulated processes, into the accessory olfactory bulb on the formation of this memory. Infusions of the calmodulin antagonist calmidazolium during the critical period prevented memory formation. However, the specific inhibitor of calcium/calmodulin-dependent protein kinase II, KN-62, or the selective inhibitor of calcium/calmodulin-dependent protein phosphatase 2B (calcineurin), FK506, was without effect on memory formation at any of the doses used. Instead of preventing memory formation, FK506 permitted the formation of a non-selective memory to strange male pheromones in the presence of mating, although FK506 alone could not induce a memory without the occurrence of mating. These results suggest that calmodulin in the accessory olfactory bulb is important in the formation of the olfactory memory to male pheromones. However, memory formation may be independent of calmodulin-kinase II. Calcineurin may play a role in processes antagonizing memory formation.     

Early onset of the rat olfactory bulb projections

Using the fluorescent carbocyanine tracer DiI, we examined in detail the early development of the projections emanating from the rat olfactory bulb. The study commenced at embryonic day 13 when the first fibres can be detected and ended at embryonic day 20, when all major fibre systems have been established. The first axons arising from the prospective olfactory bulb area are seen at embryonic day 13. Labelled fibres are provided with elaborate axonal growth cones advancing through the ventrolateral part of the telencephalic vesicle. At embryonic day 14, while the main fibre tract has not developed much further, some isolated fibres are located quite distally from the prospective olfactory bulb. These early fibres apparently course within a narrow cell-free space that extends caudally along the ventrolateral part of the telencephalic vesicle. At embryonic day 15, a number of labelled fibres form a compact bundle, corresponding to the lateral olfactory tract, that ultimately reaches the prospective primary olfactory cortex. The fibres do not stop growing, but continue to extend caudally at embryonic day 17. The results of this study provide new information on the development of axonal tracts in the olfactory system. We show that the olfactory tract projection develops earlier than the morphological appearance of the olfactory bulbs. This suggests that the early development of olfactory projections might not depend on the arrival of the olfactory epithelium axons and thus, could be governed by factors intrinsic to the neurons and/or cues present in the target environment.     

Neurocalcin immunoreactivity in the rat main olfactory bulb

The morphological characteristics and distribution of neurocalcin (NC)-immunoreactive elements were studied in the rat main olfactory bulb (OB) using a polyclonal antibody and the avidin-biotin immunoperoxidase method. NC-positive elements were abundant in the glomerular layer (GL), where numerous immunostained external tufted cells and periglomerular cells were detected. Other less abundant NC-immunolabeled populations included middle and internal tufted cells, Van Gehuchten cells, horizontal cells, vertical cells of Cajal, deep short-axon cells and granule cells. This study demonstrates the presence of NC immunoreactivity in subsets of different neuronal types in the rat main OB. This calcium-binding protein has been found in interneurons, and no evidence of immunoreactivity to NC is detected in projecting neurons. Despite the large population of labeled external tufted cells, most of them belong according to morphological criteria to the local circuit group and some others to those with interbulbar and/or intrabulbar connections. The identification of neuronal subpopulations expressing NC provides a further characterization and shows the existence of biochemical differences within morphologically identical neurons. Thus, this marker may be a useful tool in unravelling the circuitries of the rodent OB in both normal and experimental conditions. The exact physiological function of NC in the olfactory system remains unknown. On the basis of similarities to recoverin, it could be involved in mechanisms responsible for sensory adaptation. Additionally, its calcium-binding abilities may contribute to improve the temporal precision of stimuli transmission, or be concerned with general calcium-related events occurring in specific interneuronal groups.     

Identification of olfactory receptor mRNA sequences from the rat olfactory bulb glomerular layer

In situ hybridization has demonstrated mRNA for olfactory receptors (OR) in the axon terminals of olfactory receptor neurons. Neurons that express the same OR appear to send their axons to two stereotyped glomeruli in the olfactory bulb (OB). Based on these observations, we tested the feasibility of using RT-PCR to isolate and sequence OR mRNA from small samples of the rat OB glomerular layer. Biomagnetic mRNA isolation followed by RT-PCR yielded partial sequences for 21 novel members of the OR family. The results suggest that the topography of OR mRNA can be mapped across the OB, to study synaptic specificity and odor representation in the olfactory system.     

Scopolamine injection into the olfactory bulb impairs short-term olfactory memory in rats.

A previous experiment showed that systemic administration of the muscarinic antagonist scopolamine altered delayed matching in an olfactory task in rats. The present experiment tested whether the impairment could result from blockade of the cholinergic transmission in the first relay structure of the olfactory system, the olfactory bulb. 25 rats served as Ss. The drug was infused directly into both olfactory bulbs before test sessions. Results show that the intrabulbar infusion reproduced the effect of the systemic administration. With a 4-sec delay between target odor and choice test, performances of treated rats remained unchanged; however, with a 30-sec delay, rats performed randomly. Results from a complementary electrophysiological experiment in anesthetized rats support the idea that scopolamine injected into the olfactory bulb was unlikely to have reached more central structures. Further evidence for the involvement of pure sensory areas in short-term memory is concluded. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Retrograde transport of nerve growth factor from olfactory bulb to olfactory epithelium

OBJECTIVE: The purposes of this study were to determine if splenic perfusion measurements obtained using dynamic CT are useful in the evaluation of portal hypertension. MATERIALS AND METHODS: Forty-four patients with chronic liver disease (29 men and 15 women, 49-81 years old) and 38 control subjects (17 men and 21 women, 21-79 years old) underwent dynamic CT of the spleen. Regions of interest were drawn on images of the spleen and aorta, and splenic perfusion was calculated by dividing the peak gradient of the splenic time-attenuation curve by the peak aortic CT measurement increase. In 11 patients with chronic liver disease and three patients with normal livers, we measured the wedged hepatic vein pressure (WHVP) of the right or right accessory hepatic vein to estimate portal vein pressure. RESULTS: Splenic perfusion was less in patients with chronic liver disease (0.894 +/- 0.324 ml/min) than in the control group (1.299 +/- 0.429 ml/min; p < .0001). We found a significant negative correlation between splenic perfusion and WHVP (r = .741; p = .0024). CONCLUSION: A significant decrease in splenic perfusion in patients with chronic liver disease negatively correlated with WHVP. Measurement of splenic perfusion may be useful in the evaluation of portal hypertension.     

The role of the olfactory bulb in limbic mechanisms.

Reviews recent data obtained after peripheral olfactory system damage which, when compared with bulbectomy data, suggest that the olfactory bulb has certain general modulatory functions in addition to its specific sensory role as initial processor of olfactory information. It is proposed that the olfactory bulb is involved in a forebrain arousal mechanism comprised mainly of hypothalamus and limbic system. Behavioral and electrophysiological evidence is discussed that supports the idea that the olfactory bulb may modulate the excitability of forebrain regions. Anatomical details of the system are considered, and its relationship to reinforcement mechanisms and to A. Routtenberg's (see PA, Vol 42:5112 and 47:244) Arousal System II is outlined. (5 p ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Synaptic organization and neurotransmitters in the rat accessory olfactory bulb

The accessory olfactory bulb (AOB) is the first relay station in the vomeronasal system and may play a critical role in processing pheromone signals. The AOB shows similar but less distinct lamination compared with the main olfactory bulb (MOB). In this study, synaptic organization of the AOB was analyzed in slice preparations from adult rats by using both field potential and patch-clamp recordings. Stimulation of the vomeronasal nerve (VN) evoked field potentials that showed characteristic patterns in different layers of the AOB. Current source density (CSD) analysis of the field potentials revealed spatiotemporally separated loci of inward current (sinks) that represented sequential activation of different neuronal components: VN activity (period I), synaptic excitation of mitral cell apical dendrites (period II), and activation of granule cells by mitral cell basal dendrites (period III). Stimulation of the lateral olfactory tract also evoked field potentials in the AOB, which indicated antidromic activation of the mitral cells (period I and II) followed by activation of granule cells (period III). Whole cell patch recordings from mitral and granule cells of the AOB supported that mitral cells are excited by VN terminals and subsequently activate granule cells through dendrodendritic synapses. Both CSD analysis and patch recordings provided evidence that glutamate is the neurotransmitter at the vomeronasal receptor neuron; mitral cell synapses and both NMDA and non-NMDA receptors are involved. We also demonstrated electrophysiologically that reciprocal interaction between mitral and granule cells in the AOB is through the dendrodendritic reciprocal synapses. The neurotransmitter at the mitral-to-granule synapses is glutamate and at the granule-to-mitral synapse is gamma-aminobutyric acid. The synaptic interactions among receptor cell terminals, mitral cells, and granule cells in the AOB are therefore similar to those in the MOB, suggesting that processing of chemosensory information in the AOB shares similarities with that in the MOB.     

Some behavioral effects of olfactory bulb damage in the rat.

Conducted 5 experiments in which male Holtzman rats (N = 50) with either olfactory bulb or septal lesions were tested on position-habit reversal, nonappetitive passive-avoidance, 1-way avoidance, and 2-way avoidance tasks. Ss with septal damage exhibited the expected behavioral abnormalities on all tasks. Ss with bulbar damage were deficient on 1-way avoidance, were facilitated on 2-way avoidance, and could not be distinguished from the normal Ss on the other 2 tasks. (20 ref.) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Organization of inhibition in the rat olfactory bulb external plexiform layer

1. Intracellular recordings were made from the output neurons (mitral and tufted cells) of the rat olfactory bulb during electrical orthodromic stimulation of the olfactory nerve layer (ONL) and antidromic stimulation of the lateral olfactory tract and posterior piriform cortex (pPC) to test for physiological differences among the neuron types. Many of these neurons were identified by intracellular injections of biocytin, and others were identified by their pattern of antidromic activation. 2. Both marked and unmarked mitral cells showed large inhibitory postsynaptic potentials (IPSPs) in response to antidromic stimulation of the pPC, whereas tufted cells exhibited small IPSPs in response to pPC stimulation. Tufted cells, however, showed large IPSPs in response to ONL stimulation. In many cases, these tufted cell responses to ONL stimulation were larger than the mitral cell responses. The marked superficial tufted cells, those with basal dendrites in the superficial sublayer of the external plexiform layer (EPL), had the smallest IPSPs in response to pPC stimulation. These data support anatomic observations suggesting that the granule cell populations responsible for the IPSPs may be different for mitral and for superficial tufted cells. 3. The different types of output cells also showed differences in their responses to orthodromic stimulation. Type I mitral cells, which have basal dendrites confined to the deep sublayer of the EPL, were significantly less excitable by ONL stimulation than were the type II mitral cells, which have basal dendrites distributed within the intermediate sublayer of the EPL. Half of the type I mitral cells could not be excited at all by ONL stimulation. Superficial tufted cells showed even greater orthodromic excitability than type II mitral cells, usually responding to ONL stimulation with two or more spikes. 4. The ionic basis of the IPSPs in the superficial tufted cells appeared similar to those described for mitral cells. These IPSPs could be reversed by chloride injection and were associated with increased membrane conductance. 5. For both mitral and tufted cells, the number of ONL electrodes evoking IPSPs was greater than the number evoking spikes. These data suggest a kind of center-surround organization of inputs to these cells from the ONL, although this does not yet imply that the sensory receptive field of these output cells has a center-surround organization. 6. In conclusion, the properties of rat olfactory bulb output cells correlate with the sublayers of the EPL in which their basal dendrites lie.(ABSTRACT TRUNCATED AT 400 WORDS)     

Excitatory synapses in the glomerular triad of frog olfactory bulb in vitro

Whole-cell patch clamp recording techniques were applied to periglomerular (PG) cells in slices of the frog olfactory bulb (OB) to study the properties of the excitatory synapses in the triad formed by the olfactory nerve (ON) and the dendrites of mitral/tufted (MT) cells and PG cells. The postsynaptic response evoked by ON stimulation was glutamatergic and could be dissected into NMDA and non-NMDA components of equivalent amplitudes. The dendro-dendritic synapse between MT and PG cells could be activated following antidromic stimulation of the lateral and medial olfactory tract (LOT and MOT). In this case the postsynaptic potentials had amplitudes and durations comparable to those obtained by ON stimulation, the neurotransmitter was glutamate, but the synapse was largely dominated by the slow NMDA component.     

Modulation of olfactory bulb neuron potassium current by tyrosine phosphorylation

Insulin causes a suppression of whole-cell voltage-dependent outward current in cultured neurons from the rat olfactory bulb. This suppression is time-dependent; it is mimicked by application of Src tyrosine kinase inside the cell via the whole-cell patch electrode or by treatment of the olfactory bulb neurons with the tyrosine phosphatase inhibitor pervanadate. The C-type inactivation properties of the outward current in olfactory bulb neurons resemble those of the cloned Kv1.3 potassium channel. In addition, at picomolar concentrations at which it is specific for Kv1.3, the scorpion toxin margatoxin blocks most of the olfactory bulb neuron outward current. Immunocytochemical analysis demonstrates that Kv1.3 is prominent in the cultured olfactory bulb neurons. To identify specific amino acid residues that might be important for potassium current modulation, we examined the effects of pervanadate and insulin on wild-type and mutant Kv1.3 channels expressed in human embryonic kidney (HEK 293) cells. As shown previously, treatment with either pervanadate or insulin suppresses Kv1.3 current in these cells. Mutational analysis demonstrates that at least two distinct tyrosine residues are required for current suppression by pervanadate. Insulin treatment stimulates the tyrosine phosphorylation of Kv1.3 in HEK 293 cells, and a different combination of tyrosine residues is required for the current suppression by insulin. The results suggest that complex patterns of phosphorylation may be involved in the modulation of neuronal potassium current by receptor and nonreceptor tyrosine kinases.     

Are there structural and functional modules in the vertebrate olfactory bulb?

Each of twelve volunteers, at 2 week intervals, received 1 g of antipyrine, a test drug, and were exposed for 4 h either to toluene (375 mg/m3) or xylene (435 mg/m3) singly or in combination with ethanol (0.45 g/kg body wt. before the onset of exposure and 0.15 g/kg thrice every 1 h during exposure to maintain a steady level of ethanol in blood approximately 11 mmol/dm3). No significant differences were found in salivary antipyrine half-life (T1/2 approximately 12 h); and clearance (ClAP approximately 0.83 cm3/s) between control and groups exposed to solvents and/or ethanol. Nevertheless, a tendency to increase the metabolic rate of antipyrine in xylene-exposed group (T1/2 approximately 6.8 h; ClAP approximately 1.40 cm3/s) and counteraction of ethanol (T1/2 approximately 15 h; ClAP approximately 0.63 cm3/s) should be noted. The stimulation of lipid peroxidation in the serum as a biological effect of combined exposure to ethanol and toluene/xylene was observed.     

Spatial coding of odorant features in the glomerular layer of the rat olfactory bulb

Monoclonal antibodies were generated to vesicular membranes of clathrin coated vesicles enriched for acetylcholinesterase (AChE). One of these, C172, recognizes vesicles which accumulate in muscle cells around nuclei associated with acetylcholine receptor AChR clusters. Immunoblots of muscle extracts and brain purified clathrin coated vesicles show that C172 recognizes a 100 kd band in muscle, but a 180 kd band in brain. Western blots of purified AP180 protein stained with the two antibodies AP180.1 and C172 displayed the same staining pattern. Tryptic digests probed with peptide antibodies (PS26 and PS27) generated to known sequences of AP180 were used to map the epitope for C172 within the brain AP180 sequence. On immunoblots of digested AP180, all AP180 antibodies and C172 recognized a 100 kd tryptic fragment, however only C172 recognized a smaller 60 kd. Our results suggest that the C172 epitope is located within amino acids 305-598 of the AP180 sequence. Confocal fluorescence microscopy of myoblasts and myotubes stained with the C172 antibody gives a punctate immunofluorescence pattern. Myoblasts stained with C172 revealed a polarized distribution of vesicles distinct from that observed when cells are stained with gamma adaptin antibody which is known to localize to trans Golgi network. Myotubes stained with C172 antibody reveal a linear array of vesicular staining. Quantitative analysis of C172 reactive vesicles revealed a significant increase in number of vesicles present around the nuclei associated with the acetylcholine receptor clusters. These vesicles did not colocalize with the Golgi cisternae. These results indicate that a protein with homology to the neuron-specific coated vesicle protein AP180, is present in muscle cells associated with vesicles showing significant concentration around postsynaptic nuclei present in close proximity to AChR clusters.     

Glutamatergic deafferentation of olfactory bulb modulates the expression of mGluR1a mRNA

Glutamate (Glu) released by olfactory nerve axons acts on postsynaptic ionotropic and metabotropic glutamate receptors expressed by principal neurones and interneurones of the olfactory bulb (OB). Using ZnSO4 lesioning of the rat olfactory mucosa and semiquantitative RT-PCR, we examined the effect of removal of the glutamatergic input to the OB on the expression of mGluR1a, mGluR1b and GluR1 mRNAs. Two days after lesioning, mGluR1a mRNA levels in OB increased by 45%. At this time, the expression of tyrosine hydroxylase (TH) mRNA, which is strictly dependent on olfactory nerve input, was still unchanged. In contrast, 16 days after lesioning, deafferented OB exhibited a decrease in both mGluR1a (-30%) and TH (-40%) mRNAs. GluR1 and mGluR1b mRNA levels were not affected at either time point. These results suggest that alterations in glutamatergic input to OB selectively modulate the expression of the mGluR1 splicing form possessing a longer C-terminal domain.     

Chemotopic, combinatorial, and noncombinatorial odorant representations in the olfactory bulb revealed using a voltage-sensitive axon tracer

Odor information is first represented in the brain by patterns of input activity across the glomeruli of the olfactory bulb (OB). To examine how odorants are represented at this stage of olfactory processing, we labeled anterogradely the axons of olfactory receptor neurons with the voltage-sensitive dye Di8-ANEPPQ in zebrafish. The activity induced by diverse natural odorants in afferent axons and across the array of glomeruli was then recorded optically. The results show that certain subregions of the OB are preferentially activated by defined chemical odorant classes. Within these subregions, "ordinary" odorants (amino acids, bile acids, and nucleotides) induce overlapping activity patterns involving multiple glomeruli, indicating that they are represented by combinatorial activity patterns. In contrast, two putative pheromone components (prostaglandin F2alpha and 17alpha, 20beta-dihydroxy-4-pregnene-3-one-20-sulfate) each induce a single focus of activity, at least one of which comes from a single, highly specific and sensitive glomerulus. These results indicate that the OB is organized into functional subregions processing classes of odorants. Furthermore, they suggest that individual odorants can be represented by "combinatorial" or "noncombinatorial" (focal) activity patterns and that the latter may serve to process odorants triggering distinct responses such as that of pheromones.     

Stimulation of phosphoinositide hydrolysis by muscarinic receptor activation in the rat olfactory bulb

The effect of muscarinic receptor activation on phosphoinositide hydrolysis in the rat olfactory bulb was investigated by determining either the inositol (1,4,5) trisphosphate (Ins(1,4,5)P3) mass or the accumulation of [3H]inositol phosphates ([3H]InsPs). In miniprisms of rat olfactory bulb, carbachol produced an atropine-sensitive increase in Ins(1,4,5)P3 concentration. In a membrane preparation, the formation of Ins(1,4,5)P3 was stimulated by guanosine-5'-(3-O-thio) triphosphate (GTP gamma S), but not by carbachol. However, carbachol potentiated the GTP gamma S stimulation when the two agents were combined. In miniprisms prelabelled with [3H]myo-inositol, carbachol increased the accumulation of [3H]InsPs and this effect was significantly reduced by tissue treatment with either 1 microM phorbol 12-myristate 13-acetate or 1 mM dibutyryl cyclic AMP. Analysis of concentration-response curves indicated that carbachol (EC50 = 96 microM) and oxotremorine-M (EC50 = 8.2 microM) behaved like full agonists, whereas oxotremorine, BM5, arecoline and bethanechol were partial agonists. The carbachol stimulation of [3H]InsPs accumulation was counteracted with high affinity by the M1 antagonist pirenzepine (pA2 = 8.26), and less potently by the M3 antagonist para-fluorohexahydro-sila-difenidol (pA2 = 6.7) and the M2 antagonist AF-DX 116 (pA2 = 6.12). The biochemical and pharmacological properties of the muscarinic stimulation of phosphoinositide hydrolysis were compared with those displayed by the muscarinic stimulation of adenylate cyclase in the rat olfactory bulb.     

Study of glomeruli in the frog olfactory bulb using a fluorescent potential-sensitive dye

Localized irradiation of the skin and subcutaneous tissues with large single doses of gamma rays can induce immediate effects characterized by erythema, desquamation, and necrosis. Correlations between the evolution of the lesions and dosimetry studies have to be established by biophysical methods. NMR studies of the effects of an irradiated Fricke solution might be a means of controlling the delivered irradiation doses. After exposition to ionizing radiations, ferrous ions are transformed into ferric ions. Both are paramagnetic ions, and proton spin-lattice relaxation is accelerated depending on the oxidation reaction. In this study, solution of ammonium ferrous sulfate in an acid environment was incorporated into a gelling substance made with agarose, so that T1 weighted image contrast could be used to detect ferric ion formation. Experiments with 192Ir and 60Co gamma rays with doses in the 0 to 100 Gy range were conducted with Fe2+ concentrations of 0.5, 1, 1.5, and 2 mM in a gelling substance containing 4% agarose. A relationship was established between the amount of Fe3+ created and the spin-lattice proton relaxation rate, which led to a straightforward dose-effect relation. The use of such high doses allowed us to reproduce realistic conditions of accidental overexposure. A linear relationship was obtained between the doses absorbed and the NMR parameters measured (T1 and relative image intensity).     

Sodium current in periglomerular cells of rat olfactory bulb in vitro

The capacity of periglomerular cells (PGc) to give fast, Na-dependent action potentials is a crucial and debated issue for the comprehension of how sensory information is processed in the olfactory bulb (OB). Using patchclamp whole cell recording in thin slices of rat OB (P8-P20) we showed that fast sodium conductance is present in all the PGc studied, that this current is sufficiently large to generate action potentials and that action potentials can be evoked in these cells by direct stimulation of the olfactory nerve. A comprehensive kinetic characterization of INa is also presented.