Impacts of an urban flood control infrastructure on the limnology and ichthyofauna of a basaltic Cuesta stream (southeast Brazil).

Strategies for flood control associated to extreme precipitation events in urban areas are urgent, in order to prevent not only material damages but also to avoid human losses. The construction of flood contention reservoirs ("piscinões") has become a common engineering intervention in urban and peri-urban areas. However, there is a lack of studies focused on the evaluation of environmental impacts of this type of construction. This study intended to verify the ecological effects of a retention reservoir built directly on the course of the Cascata stream, Botucatu (SP). Three sampling sites were selected, located upstream the reservoir, in the reservoir and downstream. Samplings were carried out in July (winter - dry) and November (late spring - rainy) 2020. In situ measurements were obtained through a multiparameter probe (temperature, pH, electrical conductivity, dissolved oxygen, total dissolved solids, and oxidation-reduction potential) and water samples were collected for laboratory determinations (nitrogen, total phosphorus, thermotolerant coliforms, and chlorophyll-a). For fish sampling, manual trawls, sieves and hand nets were used, with a sampling effort of 10 throws per artefact and site. Despite the small distance between the sampling points (~1,300 m) considerable changes in the limnological conditions and fish community structure were observed. The studied environment is originally a small river surrounded by riparian forest, but this characteristic was abruptly changed in the reservoir stretch, with the direct exposition of a much larger water surface to intense solar radiation and atmosphere exchanges. Consequently, as evidenced by the PCA analysis, there was a considerable (stream-reservoir increase) of temperature, dissolved oxygen and chlorophyll. However, this spatial trend was partially disturbed by an accidental sewage-pipe rupture (posteriorly fixed) adjacent to the first sampling point, due to a previous event of extreme precipitation, which resulted in increased values of nutrients, chlorophyll, conductivity and thermotolerant coliforms. Eleven fish species were collected (two non-native), belonging to seven families and five orders. The upstream reference point (despite not be pristine), was characterized by the predominance of native species, while the reservoir condition favored the development of large populations of the non-native species. Despite the urgency of effective actions to prevent floods in urban areas, construction of contention reservoirs directly on stream courses should be avoided, due to their negative ecological impacts.

Gap junctions in the rat cochlea: immunohistochemical and ultrastructural analysis.

Gap junctions in the rat cochlea were investigated using immunostaining for connexin26 and transmission electron microscopy. Electron microscopy of normal and pre-embedded immunostained material showed that there were gap junctions between and among all cells that light microscopy showed to have immunostained appositions. Light microscopy showed immunostaining between and among cell types that electron microscopy showed to be joined by gap junctions. Immunostaining for connexin26 was therefore taken as providing a reasonable approximation of the locations of gap junctions throughout the cochlea and was used to provide an overview of the extent of those locations. Cells interconnected via gap junctions fell into one of two groups. The first group consists of nonsensory epithelial cells and includes interdental cells of the spiral limbus, inner sulcus cells, organ of Corti supporting cells, outer sulcus cells, and cells within the root processes of the spiral ligament. The second group consists of connective tissue cells and includes various fibrocyte types of the spiral limbus and spiral ligament, basal and intermediate cells of the stria vascularis, and mesenchymal cells which line the scala vestibuli. The present work represents a first attempt towards a description of how serial gap junctions among cochlear cells reflect a level of organization of the tissue. The organization described here, together with a great deal of information from previous investigators, suggest that serially arranged gap junctions of both epithelial and connective tissue cells serve as the structural basis for recycling endolymphatic potassium ions that pass through the sensory cells during the transduction process.

Treatment of endolymphatic hydrops with cochleosacculotomy. Clinical results and experimental findings.

Cochleosacculotomy was performed on 25 patients with Meniere's syndrome. Long-term relief of vertigo was obtained in 19 out of 23 (82%). Postoperatively dead ears occurred in three cases. The same operation was performed on 12 guinea pigs in which hydrops had been surgically induced by blockage of the endolymphatic duct and sac. All fistulas were healed and cochleosacculotomy did not decrease or prevent the induced endolymphatic hydrops in these animals. Although histologic confirmation of persistent fistulas in human ears is lacking, the relief of vertigo in patients may not be caused by "drainage" but, rather, by a nonspecific effect on the inner ear. Cochleosacculotomy gives results comparable with other nondestructive surgical procedures performed to suppress vertigo in Meniere's syndrome.

Effect of ototoxic drug administration to the endolymphatic sac.

Aminoglycosides were applied to the endolymphatic sac of guinea pigs, and the inner ears were studied by light microscopy. In group 1 gentamicin sulfate application on the lateral surface of the endolymphatic sac produced sensory cell atrophy limited to the macula sacculi. In group 2 gentamicin injection into the sac by extradural and intradural approaches often showed lesions in the maculae sacculi and to the sac from a lateral opening resulted in a similar pattern of degeneration but with less severity in comparison to the injection method. Mild to severe endolymphatic hydrops was sometimes observed. The study shows diffusion of drugs taking place against the longitudinal flow of endolymph. This procedure may be applicable for the treatment of Meniere's disease.

Morphology of the membrana limitans.

The morphology of the membrana limitans (ML) was studied by light and electron microscopy to further understand its functional significance. The ML is a porous shield composed of a few to several layers of tightly packed fibrocytes that separate the perilymphatic space of the cochlea and saccule from that of the utricle and canal system. These fibrocytes are thinner on the side of the vestibule. They show many desmosomes and gap junctions and are interspersed with fibrils. The 3-dimensional reconstruction from celloidin sections shows that the ML, as it is viewed from the anterior side, is rectangular in shape, with various surface curvatures and an invagination toward the internal aperture of the vestibular aqueduct. Horseradish peroxidase, a tracer substance, placed on either side of the ML fails to pass to the opposite side. The ML and trabecular meshwork may be a factor contributing to differential sensitivity of cochlear and vestibular sensory cells in the presence of noxious substances.

Effect of fistulae on endolymphatic hydrops.

An experiment was performed on 31 guinea pigs to study the effect of fistulae of the horizontal canal, superior canal, common crus, and utricle on the course of experimentally induced endolymphatic hydrops. The effect of fistulae on corresponding parts of normal ears was also studied in an additional 17 animals. The results indicated a remarkable consistency of healing of the fistulae in both groups of animals. Fistulae had no significant effect on the course of endolymphatic hydrops, irrespective of whether the fistulae were made immediately before or several months after obliteration of the endolymphatic duct or whether a polyethylene strut was introduced into the membranous labyrinth. Once again, the functional significance of the endolymphatic sac was clearly apparent, for ablation of the sac consistently produced severe hydrops and atrophic changes in sensory and neural structures. Assuming that the mechanisms of labyrinth repair in the guinea pig are comparable to the higher mammalian ear, it is apparent that procedures designed to surgically fistulize the membranous labyrinth are of questionable value and are probably not rational therapeutic approaches to the management of Ménière's disease.

Changes in immunostaining of cochleas with experimentally induced endolymphatic hydrops.

Cochleas with experimentally induced endolymphatic hydrops were immunostained for Na+,K(+)-ATPase, intracellular Ca(++)-ATPase, carbonic anhydrase, aldehyde dehydrogenase, calcium-binding proteins, vimentin, and the gap junction protein, connexin 26. No changes in immunostaining of hydropic ears were observed 1 week after blockage of the endolymphatic duct. Two weeks to 1 month after the operation, immunostaining of type I fibrocytes in the spiral ligament, which are positive for all but Na+,K(+)-ATPase, was slightly decreased on the operated side. These changes became more pronounced 3 months after the operation. However, staining for Na+,K(+)-ATPase of the stria vascularis and of type II fibrocytes of the spiral ligament was not reduced until 6 months postoperative. The reduction of enzymes and other cell constituents that may be involved in ion balance of cochlear fluids indicates that cells in the spiral ligament play an important role in cochlear homeostasis and that they merit further study in animal and human otopathology.

Degeneration of vestibular sensory cells caused by ablation of the vestibular aqueduct in the gerbil ear.

The vestibular aqueduct of the gerbil has a unique anatomic feature that makes it possible to selectively obliterate the endolymphatic sac with or without interfering with its venous drainage. In animals in which only the endolymphatic sac was ablated, endolymphatic hydrops was slight in the cochlea and was absent in the vestibular labyrinth. The cochlear and vestibular sensory cells were normal. In animals in which both the endolymphatic duct and the vein were obliterated, hydrops was slight, with the exception of a few cochleas that showed moderate hydrops. The sensory cells of the posterior canal cristae had degenerated in all specimens, while varied pathologic changes in cochlear and vestibular sensory cells were present in some specimens. These results suggest that hydrops is primarily due to blockage of the endolymphatic duct and sac and that degeneration of sensory cells occurs when blood flow in the vestibular aqueduct is impeded. Pathologic changes in the endolymphatic sac, including the vascular plexus at the endolymphatic sac, may play an important role in the production of endolymphatic hydrops and vestibular symptoms in Meniere's disease.

Vestibular responses of normal and hydropic ears of the guinea pig to middle ear pressure application.

The pressure in the middle ears of normal and hydropic guinea pig ears was increased, and nystagmographic recordings were compared. Two-month unilateral hydropic guinea pigs and normal control guinea pigs underwent pressure treatments in which pressure was introduced into the middle ear. Significantly lower pressure was needed to elicit nystagmus in hydropic ears (mean 1.00 psi or 70.3 cm H2O) compared with normal control ears (mean 1.27 psi or 89.3 cm H2O). All of the normal control guinea pigs showed fast phase nystagmus toward the pressure-applied side, while hydropic guinea pigs showed nystagmus toward the normal ear. The duration of nystagmus was slightly longer in hydropic animals than in normal control animals. The slow phase velocity was slightly higher in the hydropic guinea pigs. Histologic examination revealed that the vestibular sensory cells remained normal and that changes in the organ of Corti were similar between the hydropic ears with and without pressure treatment at equal survival times.

Selective lesions of the vestibular labyrinth.

In attempts to decrease gentamicin ototoxicity, the round windows of guinea pigs were covered with fat before gentamicin was given through the middle ear cavity. Although ototoxicity to the cochlea was reduced, the desired effect on the vestibular sense organs also was reduced, because of the oval window's resistance to drug penetration. In another series of guinea pigs, simple opening of the lateral semicircular canal and covering it with a pledget of Gelfoam soaked with gentamicin was an excellent method for producing a selective vestibular lesion. It is expected to reduce the function of all vestibular sense organs without impairing cochlear function.

Volumetric and dimensional analysis of the guinea pig inner ear.

The objective of this study was to provide accurate volumetric data on the fluid spaces and soft tissue in the guinea pig inner ear by measuring all histologic serial sections by means of Metamorph Imaging Software at 400x to 1,000x magnification. The total endolymph volume of the inner ear was 4.691 mm3, of which 1.501 mm3 was in the cochlea, 3.090 mm3 in the vestibular labyrinth, and 0.100 mm3 in the endolymphatic duct and sac. The total perilymph volume was 15.938 mm3, of which 8.867 mm3 was in the cochlea and 7.071 mm3 in the vestibular labyrinth. The volume of the organ of Corti per millimeter length increased toward the apex, but the volumes of the stria vascularis, spiral ligament, and spiral limbus decreased. The volume of the macula utriculi was larger than that of the macula sacculi. The measurement of the luminal surface area of the stria vascularis was 3.944 mm2, and that of the vestibular dark cells was 5.772 mm2.

Comparison of endolymph cross-sectional area measured histologically with that measured in vivo with an ionic volume marker.

In order to establish how endolymph volume is regulated, it is essential to be able to measure endolymph volume or cross-sectional area in vivo. We have developed methods to accomplish this by injecting the volume marker ion hexafluoroarsenate (AsF6) into endolymph by iontophoresis. For an injection at a constant rate, the endolymph concentration is inversely dependent on the cross-sectional area of the scala into which injection occurred. Marker concentrations were monitored by inserting ion-selective microelectrodes into endolymph near the injection site. In a previous study we quantified the degree of hydrops in animals following ablation of the endolymphatic sac. In the present study we validated the technique by comparing the endolymphatic cross-sectional area measured in vivo with AsF6 with that measured by established histologic procedures. The correlation between the two measures was good, with a coefficient of .903, although the area measured histologically was a little lower than that measured in vivo.

Electron microscopic cochlear observations in bilateral Ménière's disease.

Electron microscopic study of the cochleas of an individual with bilateral Ménière's disease revealed the presence of many abnormal sensory cells in the apical regions of the cochleas. The pathological alterations were greater in the left ear with the greater hearing loss. There were some giant cilia, fusion of cilia, and loss of cilia. The outer hair cells contained diffuse cuticular bodies near or basal to the nuclei. Many outer hair cells were retracted away from the reticular membrane. The population of the nerve endings appeared normal. The inner hair cells of the apical turns appeared essentially normal. The spiral ganglia were normal in number and morphology at the apical turns, and a majority of their cell bodies were of the unmyelinated or partly myelinated types. The stria vascularis showed atrophy; however, the magnitude of this change was consistent with that known to occur in the aging ear. In the distended areas Reissner's membrane showed areas devoid of mesothelial cells, as well as atrophic epithelial cells. The blood vessels were no different from other human cochlear vessels. These abnormalities seen in the stereocilia, the outer hair cells, and Reissner's membrane are a matter of fact. We must admit, however, that not enough electron microscopic studies have been performed on ears from aging individuals or ears with other pathologies to state that these changes are unique to Ménière's disease. Furthermore, there is currently insufficient knowledge to predict whether these changes affect auditory function.

Electron microscopic temporal bone histopathology in experimental pneumococcal meningitis.

Bacterial meningitis is one of the most common causes of acquired profound sensorineural deafness in children. Measurement of hearing and examination of the cochlea is limited in patients suffering from acute meningitis. A rabbit model of pneumococcal meningitis was developed to identify the temporal bone histopathologic changes that occur in meningogenic labyrinthitis caused by Streptococcus pneumoniae. Light microscopy was previously performed on temporal bones from acutely meningitic rabbits with profound hearing loss as determined electrophysiologically. Extensive inflammation of the cochlea with endolymphatic hydrops was observed. The organ of Corti, however, showed preserved architecture in the majority of these animals. In order to further investigate these findings, a protocol was used to create meningitic rabbits with hearing loss ranging from early high-frequency loss to profound deafness. The temporal bones from 7 rabbits were examined by transmission electron microscopy. In cases of mild hearing loss, partial degeneration of the inner row of outer hair cells, as well as edema of efferent cochlear nerve endings and marginal cells of the stria vascularis, was seen. With increasing degrees of hearing loss, the remainder of the organ of Corti and intermediate cells of the stria showed ultrastructural abnormalities. Spiral ganglion cells and basal cells of the stria vascularis remained intact in all subjects. This study provides unique information regarding the histology and pathophysiology of meningogenic deafness. The clinical implications of these findings are discussed, with an emphasis on potentially reversible changes and therapeutic intervention.

Scanning electron microscopic observations on the distended Reissner's and saccular membranes in the guinea pig.

The surface ultrastructures of membranous labyrinths in normal and hydropic guinea pig inner ears were studied. In normal specimens differences between Reissner's and saccular membranes were observed. The endolymphatic epithelial cells of the saccule were smaller in size and their microvilli varied more in size and distribution. Mesothelial cells of the saccule were often spherical in shape and bulged toward the vestibule. In hydropic guinea pig inner ears, distension of Reissner's and saccular membranes occurred by irregular enlargement of epithelial cells. Microvilli were reduced in size and number. On the severely distended Reissner's membrane, outpouchings, infoldings and holes were observed. Transmission electron microscopic examination revealed no significant difference between the tight junctions of epithelial cells in normal or hydropic Reissner's membranes. Mesothelial cells of Reissner's membrane were more severely degenerated than those of the saccule. Cochlear sensory cells were often degenerated in the apical turn where mesothelial cells were lacking, but saccular sensory cells showed very little change in their surface structures.

Computer-aided three-dimensional reconstruction and morphometry of the outer hair cells of the guinea pig cochlea.

The outer hair cells and their nerve endings in the basal and third turns of the guinea pig cochlea were reconstructed three-dimensionally from serial thin sections by means of computer graphics, and morphometric data were obtained. The number of nerve endings in the third turn was two to three times greater than that in the basal turn. Many afferent and efferent terminals in the third turn did not demonstrate synaptic specialization. Presynaptic dense bodies were missing in the majority of outer hair cells in both basal and third turns. The morphologic arrangement of the subsurface cisternae and efferent fiber synapses on the side and base of the outer hair cells suggests a close functional relationship. The nerve fibers and cisternae may be involved in the contractile process of the cells. The volume of the outer hair cells in the first row of the basal turn was about 656 micron 3, and third turn, 1358 micron 3. The total count of mitochondria in the outer hair cells of the first row in the basal turn was 1425, and 1963 in the third turn. The density of mitochondria in the sensory cell in the basal turn was higher. The highest density was seen in the infranuclear region. The mitochondrial distribution patterns suggest that metabolic activity of the outer hair cells is higher in the basal turn than in the third turn and the energy requirement is greatest in the region close to nerve endings.

Ischemia of the endolymphatic sac.

A decrease in vascular density in the endolymphatic sac was suspected as a factor in the pathogenesis of endolymphatic hydrops in Meniere's disease. The present study was undertaken to explore this possibility by cutting the posterior meningeal artery and the sigmoid sinus above and below the external aperture of the vestibular aqueduct or by incision of the dura adjacent to the sinus in 18 guinea pigs. The lesions in the sac were greater in the segmental ablation of the artery and sinus and were consistently associated with the development of endolymphatic hydrops. Among the lesions shown in the sac epithelia, the intermediate portion was most often and most severely affected with a decrease in rugose formation and a flattening of the tall epithelial cells or replacement of epithelial cells by squamous type cells. A high correlation between the lesions in the intermediate portion and occurrence of hydrops suggests that the intermediate portion plays a greater role in the pathogenesis of endolymphatic hydrops. The sac luminal precipitates known to be increased in human Meniere's cases were decreased or absent in this study, which suggests that the increased amount is unlikely to be the cause of endolymphatic hydrops. The evidence supports the hypothesis that these substances are secreted by the endolymphatic sac. The limited sensory cell lesions seen in the cochleae and saccules are likely to be due to a temporary vascular ischemia and endolymphatic hydrops.

Effect of diuretics on endolymphatic hydrops.

The effect of diuretics on endolymphatic hydrops, which was produced by obliteration of the endolymphatic duct and sac in guinea pigs, was determined by statistical analysis of volumetric changes in the scala media. When acetazolamide was administered orally at 159 to 434 mg/kg per day for a period of 30 days, endolymphatic hydrops did not reach the extent of distension shown in animal ears without drug treatment. No difference was seen in animals with one month of drug treatment followed by three additional months without treatment. This result suggests that acetazolamide has the effect of reducing endolymphatic hydrops only during the period of drug administration. On the other hand, there appeared to be no effect, statistically, on hydrops in animals which were treated with chlorthalidone 72 to 182 mg/kg every other day for one month with survival times varying from one to four months. The effect of this drug favorably reported for the treatment of Meniere's disease symptoms is not demonstrated on endolymphatic hydrops, but rather is likely to involve other mechanisms. There was no essential difference in the extent of cochlear sensory and neural atrophy in the inner ears of animals with or without these drug treatments.

Vestibular sympathetic nervous system in guinea pig.

The vestibular sympathetic fibers were examined in 20 guinea pigs by the immunohistochemical demonstration of tyrosine hydroxylase and dopamine B-hydroxylase. The vestibular sympathetics originated in the ipsilateral superior cervical ganglion and entered the internal auditory meatus along the labyrinthine artery. At the Schwann-glial border, some of the sympathetic fibers left the artery and went into the superior and inferior divisions of the vestibular nerve and formed a loose meshwork among the Scarpa's ganglion cells, while other fibers continued to follow the labyrinthine artery. Both groups of fibers entered the cristae ampullares and saccular and utricular maculas after several bifurcations in the cribrose areas and terminated either near the capillaries beneath the sensory epithelia or among the vestibular nerve fibers. These fibers traveled freely in the vestibular labyrinth without being restricted to following blood vessels or vestibular nerve fibers. Some sympathetic fibers made direct contact with the vestibular efferent fibers or the vestibular afferent fibers at the node of Ranvier. Sympathetic fibers were not observed in the sensory epithelia or semicircular canals, and were rarely found in the vicinity of the dark cells.

Cholinergic and noradrenergic nervous systems in the cynomolgus monkey cochlea.

The cholinergic and noradrenergic nervous systems in the cochleas of cynomolgus monkeys were examined by histochemical and immunohistochemical demonstration of acetylcholinesterase (AChE) and tyrosine hydroxylase (TH) and/or dopamine B-hydroxylase (DBH), respectively. AChE activity was demonstrated in the efferent fibers making synaptic contact with the perikarya and the cell processes of type II neurons. Type II neurons also demonstrated AChE activity on the cell surface and their neurites, suggesting that the AChE-positive fibers in the simian cochlear nerve trunk, previously believed to be of efferent origin, are the central processes of type II neurons. TH-like and DBH-like immunoreactivity of beaded unmyelinated fibers demonstrated similar distribution throughout the cochlear nerve trunk, Rosenthal's canal, and osseous spiral lamina. During their course, they came in contact with unmyelinated efferent fibers and neurites of type II neurons and abutted blood vessels. However, their distribution was not restricted to following blood vessels or cochlear nerve fibers. These observations indicate that the sympathetic nerve supply and olivocochlear efferent system in the cochleas of monkeys differ from those of smaller animals.