Polyhydroxyalkanoate (PHA) accumulation potential and PHA-accumulating microbial communities in various activated sludge processes of municipal wastewater treatment plants.

AIMS: To clarify the polyhydroxyalkanoate (PHA) accumulation potential and the PHA-accumulating microbial community structure in activated sludge in municipal wastewater treatment plants (WWTPs) and to identify their influential factors. METHODS AND RESULTS: Nine activated sludge samples were collected from municipal WWTPs employing various biological treatment processes. In acetate-fed 24-h batch experiments under aerobic and nitrogen- and phosphorus-limited conditions, polyhydroxybutyrate (PHB) content of activated sludge increased from 0-1·3 wt% to 7·9-24 wt%, with PHB yields of 0·22-0·50 C-mol 3-hydroxybutyrate (C-mol acetate)(-1). Microbial community analyses found that activated sludge samples that accumulated >20 wt% of PHB after 24-h PHA accumulation experiments had >5·0 × 10(8) copies g(-1)-mixed liquor-suspended solid of phaC genes. CONCLUSIONS: Results indicated that (i) activated sludge in municipal WWTPs can accumulate up to approx. 20 wt% of PHA without enrichment processes, (ii) PHA accumulation potential of activated sludge varied depending on the operational conditions (treatment processes) of WWTPs, and (iii) phaC gene number can provide a simple indication of PHA accumulation potential. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first study to compare the PHA accumulation potential and PHA-accumulating microbial communities in activated sludge of various treatment processes. Our findings may be useful for enhancing the resource recovery potential of wastewater treatment systems.

Effects of locus coeruleus stimulation on neuronal activities of dorsal lateral geniculate nucleus and perigeniculate reticular nucleus of the rat.

In rats anesthetized with urethane, a stimulating electrode was introduced to the locus coeruleus by observing the antidromic field response to single shock stimulation of the dorsal pathway of noradrenergic axons. Effects of locus coeruleus stimulation were studied on activities of relay neurons and intrinsic interneurons of the dorsal lateral geniculate nucleus and on those of neurons in the perigeniculate reticular nucleus. The intrinsic interneurons and the perigeniculate reticular neurons are believed to exert inhibition upon the relay neurons. The relay neurons were activated by repetitive stimulation of locus coeruleus; spontaneous discharges were increased in rate and the threshold of response to single shock stimulation of the optic nerve was lowered. The activation was rarely seen in rats pretreated with alpha-methyl-p-tyrosine. Iontophoretic application of phentolamine, an alpha-blocker, effectively antagonized the activation, whereas an iontophoretic beta-blocker and cholinergic blockers were virtually ineffective. The activation of the relay neurons was suggested to be due to a direct action of noradrenaline, released by locus coeruleus stimulation. Locus coeruleus stimulation inhibited the interneurons and activated the perigeniculate reticular neurons; spontaneous or light-evoked discharges were suppressed in the interneurons and tonic discharges were elicited in the perigeniculate reticular neurons. These effects of locus coeruleus stimulation were mimicked by noradrenaline applied iontophoretically. Activation of the perigeniculate reticular neurons was antagonized by an iontophoretic alpha-blocker but not by a beta-blocker. Two special features emerge from the present results: (1) the locus coeruleus exerts different effects upon the two neuronal constituents of the dorsal lateral geniculate nucleus, excitation of the relay neurons and inhibition of the intrinsic interneurons; (2) a suggestion previously advocated that locus-coeruleus-induced excitation of the lateral geniculate relay neurons would be due to inhibition of inhibitory neurons (disinhibition) does not hold true, at least with respect to the perigeniculate reticular neurons; the latter neurons have been proved to exert a powerful inhibition upon the geniculate relay neurons and they are excited by stimulation of the locus coeruleus.

Neuronal organization of rat thalamus for processing information of vibrissal movements.

Vibrissa-responding neurons were searched for in the somatosensory part of the thalamic reticular nucleus (S-TR) and in the ventrobasal nucleus (VB) in urethane-anesthetized rats. More than 90% of the recorded neurons of both species had receptive fields (RFs) on single vibrissae. Movements of RF-vibrissae produced a burst of multiple discharges in S-TR neurons and single spike discharges followed by a prominent suppression of spontaneous discharges in VB neurons. Antidromic invasion from stimulation of the somatosensory cortex in VB neurons was suppressed after RF-vibrissae were stimulated. A possible functional organization comprising VB and S-TR neurons for processing impulses of vibrissal movements was suggested.

Two pathways from the facial skin to the superior colliculus in the rat.

In response to single shock stimulation of the facial skin, the field potential of two negative waves (N1 and N2) appear in the intermediate layers of the rat superior colliculus (SC). From the experiments of functional ablations of the cortical facial area, the N1 wave was ascribed to the post-synaptic activity mediated by the direct trigemino-tectal pathway whereas the N2 wave to that mediated by the trigemino-cortico-tectal pathway. Single unit recordings also confirmed the existence of two pathways from the facial skin to the intermediate layers of SC.

GABA sensitivity of neurons of the visual layer in the rat superior colliculus.

Inhibitory action of iontophoretically applied GABA was examined on neurons in the visual layer of the rat superior colliculus (SC). Spontaneous discharges of all neurons tested were readily abolished by GABA ejected with currents less than 25 nA. In some cells the discharges evoked by near threshold electrical stimulation of the optic nerve or those evoked by a spot of light moving across receptive fields were suppressed by the same dose of GABA as that required to abolish the spontaneous discharge. However, in other cells the evoked discharges were much more resistant to GABA than the spontaneous activity. GABA sensitivity of the evoked activities was examined on various classes of SC cells which were identified by their recording depth, response latency to electrical stimulation of the optic chiasm and other properties. SC cells of the visual layer were classified into 8 types: classes Ia and Ib in the most superficial layer (N3 zone), class II in the thin layer below the N3 (N2 zone) and classes IIIa, IIIb, IVb and IVc in the deepest layer below the N2 (N1 zone). Effects of GABA upon these cell classes are summarized as follows; (1) Ia and IVb cells were readily suppressed by GABA, (2) Ib and II and most of IIIa and IVc cells were GABA-insensitive, and (3) GABA sensitivity varied from cell to cell in classes IIIb and IVa.

Inhibition of neuronal activity of the substantia nigra by noxious stimuli and its modification by the caudate nucleus.

In urethane-anesthetized rats discharges of neurons of substantia nigra, pars compacta (SNC) were recorded extracellularly after natural somatic sensory stimulation and electrical stimulation of peripheral sensory nerves. (1) Among different modalities of somatic sensory stimulation tested, noxious stimuli were effective in reducing spontaneous discharges of SNC neurons. The inhibition appeared with a concomitant increase of spike amplitude. The same inhibitory effect was obtained by stimulating the sciatic nerve (SC) repetitively. In response to single shock stimulation of the SC the inhibition occurred at an average latency of 39.6 msec (S.E. 1.6 msec) and lasted for 221.6 msec on average (S.E. 10.8 msec). (2) The SC-induced inhibition of SNC neurons failed to reliably block ortho- and antidromic discharges evoked from the caudate nucleus (Cd). (3) In rats with the Cd lesioned the SC-induced inhibition was longer lasting than in controls. When the Cd was stimulated concurrently with SC stimulation, the inhibition from the SC was weakened. (4) In a majority of SNC neurons, their inhibition by SC stimulation, their inhibition by SC stimulation was antagonized by intravenous injection of haloperidol.

Sensitivity to GABA of neurons of the dorsal and ventral lateral geniculate nuclei in the rat.

GABA was applied iontophoretically to dorsal and ventral lateral geniculate (LGd and LGv) neurons in rats. Spontaneous discharges were readily suppressed in both species of neurons. While in LGd neurons, evoked discharges by optic nerve stimulation were suppressed as readily as were spontaneous discharges, LGv neurons were characterized in that evoked discharges were much more resistant than spontaneous discharges.

Properties of ipsilateral retinogeniculate afferents in albino and hooded rats.

By recording single unit activities from the dorsal lateral geniculate nucleus in albino and hooded rats, physiological properties of the ipsilateral retinogeniculate afferents were compared with those of the contralateral ones. The results show that the ipsilateral retinogeniculate pathway was characterized by intermediate conduction velocities, relatively high incidence of the tonic response and the visual field representation of central 30 degrees from the vertical midline on both sides.

Visual receptive-field properties of single cells in the rat superior colliculus.

Receptive-field properties were studied in single cells in the visual layer of the albino rat's superior colliculus. All the samples (N = 195) were identified as one of the four classes (I-, II-, III- and IV-cells) which were established previously on the bases of the response properties to single shocks to the optic pathway and of the recording sites. The four classes were located more deeply in the superior colliculus roughly in the order of from I to IV. Response latencies to single shocks to the optic chiasm were shortest in the III-cells, intermediate in the II-cells and longest in the I- and IV-cells. These cell classes turned out to be different from each other in a number of visual properties. 1) Receptive-field centers in the I- and II-cells (N = 26 and 27, respectively) were of either the ON-OFF or the ON type. Of class III (N = 47), one subgroup (IIIa) consisted mostly of the OFF type whereas the other (IIb) was made up mostly of the ON-OFF type. About two-thirds of the IV-cells (71/95) were of either the ON, the OFF or the ON-OFF type; except for two cells, the rest were all of the moving-sensitive type. 2) The average size of the receptive-field center was small (5.4 degrees +/- 2.3 degrees) in the class I, intermediate in the classes II and III (8.1 degrees +/- 2.1 degrees and 7.1 degrees +/- 2.7 degrees, respectively) and large in the class IV (18.1 degrees +/- 15.9). 3) Responsiveness to moving light stimuli (speeds, 40 degrees--90 degrees/sec) decreased in the order of IIIa, II and IIIb, and I and IV. 4) Maintained activity was lower in the superficially located cells (I, II and IIIa) than in the deeply located ones (IIIb and IV).

Receptive-field properties of cells in the dorsal part of the albino rat's lateral geniculate nucleus.

Receptive-field properties of 273 relay (principal, P-) cells of the dorsal lateral geniculate nucleus (LGd) were studied in urethane-anesthetized albino rats, in an attempt to see if there is some relation between the visual property and the conduction velocity of afferent optic nerve fibers. According to properties of the receptive-field center, P-cells were classified into two types, common (89%) and uncommon (11%). The common type consists of OFF-phasic, ON-phasic, ON-tonic and ON-OFF-phasic cells, while the uncommon type includes ON-inhibited, moving-sensitive, ON-OFF-inhibited, simple-cell-like and complex-cell-like cells. The mean response latency to single optic chiasm shocks increases in the order of OFF-phasic (1.94 msec), ON-phasic (2.35 msec), ON-tonic (2.87 msec), ON-OFF-phasic cells (3.04 msec) and uncommon type (3.18 msec). The mean size of the receptive-field center in each of the four common types was smaller than that in the uncommon type; 6 degrees--7 degrees vs. 11 degrees. From responsiveness to moving light spots with speeds faster than 25 degrees--30 degrees/sec, P-cells of the common type were divided into the fast- and slow-movement-sensitive cells. The ratio of occurrences of fast- to slow-movement-sensitive cells decreases in the order of the OFF-phasic (2.7), ON-phasic (2.4), ON-tonic (1.1) and ON-OFF-phasic types (0.06). The optic chiasm latencies were shorter than 2.5 msec in most of the fast-movement-sensitive cells while the reverse was true for most of the slow-movement-sensitive cells. From these findings discussions were made to point out that the rat LGd mainly consists of Y- and W-like P-cells and that the Y/W dichotomy of P-cells approximately corresponds to the previously established fast/slow classification.

Characteristics of optic nerve innervation in the rat superior colliculus as revealed by field potential analysis.

With lightly anesthetized rats, field responses to single optic nerve (ON) and optic chiasm shocks were studied in superficial layers of the superior colliculus, i.e., St. zonale (SZ), St. griseum superficiale (SGS) and St. opticum (SO). Based upon characteristics in the depth profile of the field response within the superficial layers, we identified three zones of N3, N2, and N1 from the SZ in this order. The narrow N2-zone was localized around the middle part of the SGS and less distinct than the other two zones. The N3-zone was characterized by a N3-wave and the N1-zone with N1- and P3-waves. N3- and P3-waves were activated by slowly conducting ON fibers (5.0 +/- 1.4 and 4.2 +/- 0.6 m/sec, respectively) while the N1-wave was by fastest conducting fibers (16.3 +/- 4.1 m/sec). According to recent Golgi studies, hypothetical mechanisms underlying the three waves were proposed: the N3-wave was ascribed to the activities of small vertical fusiform cells in the SZ or the upper half of SGS, the N1-wave to those of narrow field vertical cells which predominate in the lower half of SGS, and the P3-wave to those of wide field vertical cells whose somata are in the lower half of SGS or in the SO. The N3-, N1-, and P3-waves were found to differ from each other in recovery function tested with double ON shocks as well as in the topographical analysis of amplitudes through a whole extent of the colliculus. The three systems represented by the three waves were suggested to play different roles in visual information processing within the superficial strata.

A four group classification of the rat superior collicular cells responding to optic nerve stimulation.

With lightly anesthetized rats, activities of single cells responding to single optic nerve (ON) shocks were recorded from the superficial layers of the superior colliculus, i.e., St. zonale(SZ), St. griseum superficiale (SGS) and St. opticum (SO). According to response latencies and recording depths, four classes of cells were identified: I-, II-, III-, and IV-cells. Class I cells, recorded in the SZ or the upper half of SGS, were innervated by slowly conducting ON fibers of velocities slower than 4 m/sec. Class II cells, recorded from the middle part of the SGS, were innervated by ON fibers of intermediate velocity (4--8 m/sec). Class III cells, recorded from the lower half of SGS or the upper part of SO, were innervated by fast conducting ON fibers (faster than 8 m/sec). Class IV cells were recorded from the depth similar to the class III but they were innervated by slowly conducting fibers (4-1.7 m/sec). Class I cells revealed a long-lasting inhibition after initial excitation, being a sharp contrast with cells of classes II and III which were almost free from inhibition. Inhibition exerted upon class IV cells were of intermediate strength. Possible anatomical correlates of the four classes of cells and hypothetical modes of their ON innervation were discussed.

Cortically induced inhibition of neurons of rat substantia nigra (pars compacta).

The effects of electrical stimulation of prefrontal cortex upon neurons of substantia nigra (pars compacta) in anesthetized rats were mostly inhibition without antidromic excitation. By studying nigral neurons in which the inhibition from caudate-putamen was antagonized by iontophoretic bicuculline, it was found in only half of them that the same drug also antagonized the inhibition from prefrontal cortex.

Liquid chromatographic determination of oxytetracycline in swine tissues.

A simple and rapid method was developed for determination of oxytetracycline (OTC) in swine muscle and kidney by liquid chromatography (LC). The method involved homogenization of sample in acetonitrile-1M imidazole buffer containing 10 mM disodium ethylenediaminetetraacetic acid (Na2.EDTA) and 50 mM magnesium acetate (15 + 85) with added hexane, centrifugation, removal of the hexane phase, and ultrafiltration of the supernatant. L-column ODS (150 x 4.6 mm) with a mobile phase of acetonitrile-1M imidazole buffer containing 50 mM magnesium acetate and 10 mM Na2.EDTA (10 + 90) was used for the LC separation. A fluorescence detector was used at an excitation wavelength of 380 nm and an emission wavelength of 520 nm. The calibration graph was linear from 1.25 to 200 ng OTC. Recoveries of OTC from swine tissue fortified at levels of 0.05-1.0 microgram/g ranged from 58.0 to 67.3%. The quantitation and detection limits were 0.05 and 0.04 microgram/g, respectively.