Transient Cochlear Ischemia Research Articles

Prevention of ischemia-induced hearing loss by intravenous administration of hydrogen-rich saline in gerbil.

Hydrogen-rich water, which is a potent antioxidant agent, was investigated for its protective effects against ischemic damage of the cochlea in gerbils. The animals were subjected to transient cochlear ischemia by occluding the bilateral vertebral arteries for l5min. Five milliliters of hydrogen-rich saline was then intravenously administered immediately after the insult. Saline without hydrogen was used as a control. Effects of hydrogen were evaluated using the auditory brainstem response (ABR) and histological studies of the inner ear. In non-ischemia animals, ABR thresholds and histological findings of the cochlea did not change by administration of saline or hydrogen-rich saline. In contrast, transient cochlear ischemia caused a 24.2±3.8dB increase in the ABR threshold at 8kHz, and a decrease of 14.1%±1.8% in the number of inner hair cells (IHCs) at the basal turn on day 7. Ischemic damage was more severe at 16 and 32kHz. When the animals were treated with hydrogen-rich saline, cochlear damage was significantly reduced: the increase in ABR threshold was 11.7±2.6dB at 8kHz and the IHC loss was 7.5%±2.1% at the basal turn on day 7. The effects of hydrogen-rich saline were more prominent at higher frequencies. Intravenous administration of hydrogen-rich saline was effective in preventing acute hearing loss due to transient cochlear ischemia.

Intravenous administration of bone marrow mononuclear cells alleviates hearing loss after transient cochlear ischemia through paracrine effects

Bone marrow mononuclear cells (BMMCs) are known to enhance recovery from ischemic insults by secreting angiogenic factors and inducing the expression of angiogenic factors from host tissues. Therefore, the transplantation of BMMCs is considered a potential approach to promoting the repair of ischemic damaged organs. Here, we investigated the influence of BMMCs on progressive hair cell degeneration after transient cochlear ischemia in gerbils. Transient cochlear ischemia was produced by extracranial occlusion of the bilateral vertebral arteries immediately before their entry into the transverse foramen of the cervical vertebra. An intravenous injection of BMMCs prevented ischemia-induced hair cell degeneration and ameliorated hearing impairment. A tracking study showed that BMMCs injected into the femoral vein were limited in the spiral artery of the cochlea, suggesting that, although transplanted BMMCs were retained within the spiral ganglion area of the cochlea, they were neither transdifferentiated into cochlear cells nor fused with the injured hair cells and supporting cells in the organ of Corti to restore their functions. We also showed that the protein level of neurotrophin-3 and glial cell line-derived neurotrophic factor in the organ of Corti was upregulated after treatment with BMMCs. These results suggested that BMMCs have therapeutic potential possibly through paracrine effects. Thus, we propose the use of BMMCs as a potential new therapeutic strategy for hearing loss.

Liposome-encapsulated hemoglobin alleviates hearing loss after transient cochlear ischemia: An experimental study in the gerbil

The effects of liposome-encapsulated hemoglobin (LEH), an artificial oxygen carrier, were experimentally investigated in gerbils in the context of alleviation of hearing loss after transient cochlear ischemia. Animals were randomly assigned to receive 2mL/kg of either LEH (P50O2=15mmHg) or saline 1h after the experimental induction of 15min of ischemia. Sequential recordings of auditory brainstem response (ABR) showed that administration of LEH prevented hearing loss due to cochlear ischemia. The mean ABR threshold at 32kHz on day 1 was 21±7dB in the LEH group (n=6) and 45±6dB in the saline group (n=6). Thereafter, hearing impairment gradually improved up to day 7 in both groups. The animals were then subjected to histological study, which revealed that there was more substantial loss of the inner hair cells, but not the outer hair cells, in the saline group as compared to the LEH group. These results suggest that LEH is an efficient agent with regard to protection against hearing loss and underlying hair cell damager due to ischemic insult.

Experimental study of transient cochlear ischemia as a cause of sudden deafness

Experimental study of transient cochlear ischemia as a cause of sudden deafness

Protection Against Ischemic Cochlear Damage by Intratympanic Administration of AM-111

AM-111, a cell-permeable peptide inhibitor of c-Jun N-terminal kinase, was investigated for its protective effects against ischemic damage of the cochlea in gerbils. Transient cochlear ischemia was introduced in animals by occluding the bilateral vertebral arteries for l5 minutes. Then, 10 μl of AM-111 at a concentration of l, 10, or 100 μM in hyaluronic acid gel formulation was applied onto the round window 30 minutes after the insult. Gel without active substance was used in a control group. Treatment effects were evaluated by auditory brainstem response (ABR) and histology of the inner ear. In controls, transient cochlear ischemia caused a 25.0 ± 5.0 dB increase in the ABR threshold at 8 kHz and a decrease of 13.3 ± 2.3% in inner hair cells at the basal turn on Day 7. Ischemic damage was mild at 2 and 4 kHz. When the animals were treated with AM-111 at 100 μM, cochlear damage was significantly reduced: the increase in ABR threshold was 3.3 ± 2.4 dB at 8 kHz, and the inner hair cell loss was 3.1 ± 0.6% at the basal turn on Day 7. The effects of AM-111 were concentration dependent: 100 μM was more effective than 1 or 10 μM. Direct application of AM-111 in gel formulation on the round window was effective in preventing acute hearing loss because of transient cochlear ischemia.

Liposome‐Encapsulated Hemoglobin Alleviates Hearing Loss After Transient Cochlear Ischemia and Reperfusion in the Gerbil

To test liposome-encapsulated hemoglobin (LEH) in transient cochlear ischemia/reperfusion as a model of sudden deafness, Mongolian gerbils were randomly assigned to receive 2 mL/kg of either low-affinity LEH (l-LEH, P₅₀0₂ = 40 mm Hg), high-affinity LEH (h-LEH, P₅₀0₂ = 10 mm Hg), homologous red blood cells (RBCs), or saline (each group n = 6) 30 min before 15-min occlusion of the bilateral vertebral arteries and reperfusion. Sequential changes in hearing were assessed by auditory brain response 1, 4, and 7 days after ischemia/reperfusion, when the animals were sacrificed for pathological studies. h-LEH was significantly more protective than l-LEH in suppressing hearing loss, in contrast to RBC or saline treatment, at 8, 16, and 32 kHz, where hearing loss was most severe (P < 0.05 between any two groups) on the first day after cochlear ischemia/reperfusion. Thereafter, hearing loss improved gradually in all groups, with a significant difference among groups up to 7 days, when morphological studies revealed that the inner hair cells but not the outer hair cells, were significantly lost in the groups in the same order. The results suggest that pretreatment with h-LEH is significantly more protective than l-LEH in mitigating hearing loss and underlying pathological damage, in contrast to transfusion or saline infusion 7 days after transient cochlear ischemia/reperfusion.

Transient ischemia/hypoxia enhances gentamicin ototoxicity via caspase-dependent cell death pathway

Aminoglycoside ototoxicity is a common cause of drug-induced hearing loss. Toxicity is dose related, but some patients may still develop hearing loss even under safe dosage. Apart for genetic idiosyncrasy, indirect evidences imply that ischemia may increase the aminoglycoside ototoxic sensitivity because common clinical situations associated with cochlear ischemia such as noise, sepsis, and shock are known to augment the development of aminoglycoside ototoxicity. At present, a direct interaction of cochlear ischemia and aminoglycoside ototoxicity is still lacking. This study demonstrated a direct evidence of increased gentamicin (GM) ototoxic sensitivity in chronic guinea pig models of transient cochlear ischemia. No permanent auditory changes were observed after a single dose of GM (125 mg/kg) or after transient cochlear ischemia for 30 min. Persistent and significant auditory threshold shift was detected when GM was given after transient cochlear ischemia. Cochlear hair cells and spiral ganglion neurons are the major regions affected. Apoptosis contributes to hair cell death during acute interaction of ischemia and GM ototoxicity. Increased apoptotic cell death was also depicted when GM crossreacted with hypoxia in vitro, using cochlear cell lines. Generation of reactive oxygen species, loss of mitochondrial membrane potential, calcium release, and caspase-dependent apoptotic cell death were shown during the interaction of hypoxia and GM ototoxicity in vitro. This synergistic ototoxicity may be critical to aminoglycoside-induced hearing loss in clinical scenarios. The results should improve our understanding of the interacting mechanism and potential preventive strategy to aminoglycoside ototoxicity.

Ischemic tolerance in the cochlea

Ischemic tolerance in the cochlea was investigated in a gerbil model of cochlear ischemia. Transient cochlear ischemia was produced by extracranial occlusion of the bilateral vertebral arteries. The gerbils were divided into two groups; single ischemia group and double ischemia group. In the single ischemia group, animals were subjected to lethal cochlear ischemia for 15min. In the double ischemia group, animals were subjected to sublethal cochlear ischemia for 2min at 2 days before lethal ischemia for 15min. Consequently, sublethal ischemia prevented lethal ischemia-induced hair cell degeneration and ameliorated hearing impairment, suggesting ischemic tolerance in the cochlea.

Hematopoietic stem cells prevent hair cell death after transient cochlear ischemia through paracrine effects

Transplantation of hematopoietic stem cells (HSCs) is regarded to be a potential approach for promoting repair of damaged organs. Here, we investigated the influence of hematopoietic stem cells on progressive hair cell degeneration after transient cochlear ischemia in gerbils. Transient cochlear ischemia was produced by extracranial occlusion of the bilateral vertebral arteries just before their entry into the transverse foramen of the cervical vertebra. Intrascalar injection of HSCs prevented ischemia-induced hair cell degeneration and ameliorated hearing impairment. We also showed that the protein level of glial cell line-derived neurotrophic factor (GDNF) in the organ of Corti was upregulated after cochlear ischemia and that treatment with HSCs augmented this ischemia-induced upregulation of GDNF. A tracking study revealed that HSCs injected into the cochlea were retained in the perilymphatic space of the cochlea, although they neither transdifferentiated into cochlear cell types nor fused with the injured hair cells after ischemia, suggesting that HSCs had therapeutic potential possibly through paracrine effects. Thus, we propose HSCs as a potential new therapeutic strategy for hearing loss.

Ginsenoside Rb1 protects against damage to the spiral ganglion cells after cochlear ischemia

The effects of transient cochlear ischemia on spiral ganglion cells (SGCs) were studied in Mongolian gerbils. Ischemic insult was induced by occluding the bilateral vertebral arteries of gerbils for 15 min. Seven days after ischemia, the percentage of SGCs decreased to 67.5% from the preischemic baseline in the basal turn. Evaluation with immunohistochemical staining showed TUNEL-positive reactions in the SGCs with fragmented nuclei. In addition, we investigated the protective effects of ginsenoside Rb1 (gRb1) against ischemic injury to SGCs. Seven days after ischemia, the auditory brainstem response threshold shift was significantly reduced and the percentage of SGCs decreased to 90.2% from the preischemic baseline in the basal turn in the gRb1-treated group. These findings suggest that gRb1 prevented hearing loss caused by ischemic injury to SGCs in Mongolian gerbils.

Protective Effects of Glucocorticoids on Ischemia???Reperfusion Injury of Outer Hair Cells

This animal study aimed to investigate effects of glucocorticoids on the functional recovery of outer hair cells (OHCs) after transient ischemia. Distortion-product otoacoustic emission (DPOAE) was examined before, during, and after transient cochlear ischemia of 30 minutes using albino guinea pigs. DPOAE decreased to noise level during ischemia. On recirculation, DPOAE initially recovered with time until 20 minutes after the onset of reperfusion, but thereafter, the DPOAE level gradually decreased toward the noise level in the control animals. Prednisolone and methylprednisolone significantly improved the DPOAE level 60 minutes after the onset of reperfusion. The present findings suggest that glucocorticoids possess protective effects against ischemia-reperfusion injury of OHCs.

Ischemic damage increases nitric oxide production via inducible nitric oxide synthase in the cochlea

The present study was designed to elucidate the dynamic changes of nitric oxide (NO) production in the perilymph and to investigate the immunostaining for inducible nitric oxide synthase (iNOS) in the cochlea for 7 days after transient cochlear ischemia. Moreover, aminoguanidine, which is a selective iNOS inhibitor, was administrated immediately following ischemia and every 24 h thereafter for 7 days to investigate whether the production of NO is dependent on the iNOS pathway. Significant increases in the oxidative NO metabolites, nitrite (NO 2 −) and nitrate (NO 3 −), were measured on day 1 using an in vivo microdialysis and on-line high performance liquid chromatography (HPLC) system. The immunostaining for iNOS was strongly expressed on days 1 and 4 and returned to normal on day 7 after the ischemia. The administration of aminoguanidine reduced the oxidative NO metabolites on day 1 and suppressed the expression of iNOS. These findings suggest that transient ischemia causes a remarkable increase in NO production in the perilymph, which might be attributable to the iNOS pathway.

Transient cochlear ischemia and its effects on the stria vascularis

The effects of transient cochlear ischemia on the stria vascularis were studied. Fifteen minutes of ischemia decreased the endocochlear potential by up to 17.5 mV on day 1; it returned to normal on day 7. Immunostaining for Na+,K+-ATPase, a marker for the Na+/K+-pump, and for connexin 26, a marker for gap junctions, was inhibited on days 1 and 4, and returned to normal on day 7. Electron microscopy showed expansion of the intercellular space with abundant vacuolar formation in the stria vascularis. These morphological changes disappeared completely by day 7. The results indicate that transient ischemia causes a reversible functional disorder of the stria vascularis with fine structural changes, which may be owing to dysfunction of Na+/K+-pump or gap junctions.

AMPA/kainate-type glutamate receptor antagonist reduces progressive inner hair cell loss after transient cochlear ischemia

We investigated the effect of glutamate receptor antagonists on progressive inner hair cell (IHC) loss following transient cochlear ischemia in gerbils. Transient cochlear ischemia was induced by 15-min bilateral vertebral artery occlusion. An alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA)/kainate-type glutamate receptor antagonist, 6-7-dinitroquinoxaline-2,3-dione (DNQX), or an N-methyl- d-aspartate (NMDA)-type receptor antagonist, MK-801, was administered 10 min before the ischemic insult. Hearing was assessed by sequentially recording compound action potentials (CAPs) before, during, and after the ischemia. The degree of hair cell loss in the organ of Corti was evaluated in specimens stained with rhodamine–phalloidin and Hoechst 33342. On the seventh day after ischemia, the increases in the CAP threshold and the progressive IHC loss were significantly reduced in cochleae treated with DNQX, while MK-801 was ineffective. These results suggest that the AMPA receptor plays a critical role in the development of the progressive IHC loss induced by ischemia/reperfusion injury in the cochlea.

Glucocorticoids and dehydroepiandrosterone sulfate ameliorate ischemia-induced injury of the cochlea

This study aimed to evaluate the effects of steroidal drugs on the functional recovery of the cochlea after transient ischemia. Albino guinea pigs were subjected to transient cochlear ischemia of 30 min duration, and the threshold shifts of the compound action potential (CAP) from the pre-ischemic values were evaluated 4 h after ischemia. Pre-ischemic administration of a glucocorticoid, prednisolone or methylprednisolone, significantly ameliorated the post-ischemic CAP threshold shifts as compared with control animals at a relatively wide range of doses. Post-ischemic administration of these glucocorticoids also exhibited protective effects. Pre-ischemic administration of dehydroepiandrosterone sulfate significantly decreased the post-ischemic CAP threshold shifts 4 h after ischemia. The present results indicate that glucocorticoids and dehydroepiandrosterone sulfate possess therapeutic effects against ischemic injury of the cochlea, such as idiopathic sudden sensorineural hearing loss.

Adenovirus-mediated overexpression of a gene prevents hearing loss and progressive inner hair cell loss after transient cochlear ischemia in gerbils.

The use of adenoviral vectors has recently provided a novel strategy for direct gene transfer into the cochlea. In this study, we assessed the utility of an adenoviral vector expressing glial-cell-derived neurotrophic factor (GDNF) in ischemia-reperfusion injury of the gerbil cochlea. The vector was injected through the round window 4 days before ischemic insult. The distribution of a reporter transgene was confirmed throughout the cochlea from the basal to the apical turn and Western blot analysis indicated significant upregulation of GDNF protein 11 days following virus inoculation. Hearing ability was assessed by sequentially recording compound action potentials (CAP), and the degree of hair cell loss in the organ of Corti was evaluated in specimens stained with rhodamine-phalloidin and Hoechst 33342. On the seventh day of ischemia, the CAP threshold shift and inner hair cell loss were remarkably suppressed in the Ad-GDNF group compared with the control group. These results suggest that adenovirus-mediated overexpression of GDNF is useful for protection against hair cell damage, which otherwise eventually occurs after transient ischemia of the cochlea.

Transient cochlear ischemia causes delayed cell death in the organ of Corti: an experimental study in gerbils.

To elucidate whether ischemia-reperfusion can cause delayed cell death in the cochlea, the effects of transient cochlear ischemia on hearing and on neuronal structures in the cochlea were studied in Mongolian gerbils. Ischemia was induced by bilaterally occluding the vertebral arteries for 5 minutes in gerbils, which lack posterior cerebral communicating arteries. In gerbils, the labyrinthine arteries are fed solely by the vertebral arteries. Occlusion of the vertebral arteries caused a remarkable increase in the threshold of compound action potentials (CAPs), which recovered over the following day. However, 7 days after the onset of reperfusion, the threshold began to increase again. Morphologic changes in the hair cell stereocilia were revealed by electron microscopy. The number of nuclear collapses was counted in cells stained for DNA and F-actin to evaluate the degree of cell death in the organ of Corti. Changes in spiral ganglion cell (SGC) neuron number were detected, whether or not progressive neuronal death occurred in the SGC. These studies showed that sporadic fusion of hair cells and the disappearance of hair cell stereocilia did not begin until 4 days after ischemia. On subsequent days, the loss of hair cells, especially inner hair cells (IHCs), and the degeneration of SGC neurons became apparent. Ten days after ischemia, the mean percentage cell loss of IHCs was 6.4% in the basal turn, 6.4% in the second turn, and 0.8% in the apical turn, respectively, and the number of SGC neurons had decreased to 89% of preischemic status. These results indicate that transient ischemia causes delayed hearing loss and cell death in the cochlea by day 7 after ischemia.

Hypothermia prevents hearing loss and progressive hair cell loss after transient cochlear ischemia in gerbils

The effects of hypothermia on ischemia–reperfusion injury of the cochlea were studied in gerbils. Hearing was assessed by sequentially recording compound action potentials before, during and after the ischemia. The degree of hair cell loss in the organ of Corti was evaluated in specimens stained with rhodamine–phalloidin and the dye Hoechst 33342. Ischemic insult was applied to the animals by occluding the bilateral vertebral arteries for 15 min under normothermic or hypothermic (rectal temperature 32°C) conditions. Interruption of the blood supply to the cochlea caused a tremendous increase in the compound action potential threshold, which usually recovered to some extent with reperfusion. In the ischemia/normothermic group, the threshold did not return to the pre-ischemic level. The average increase in the threshold seven days after ischemia was 20.0 dB. Histologically, the hair cell loss increased gradually until four days after the ischemic insult. On the seventh day, the mean loss of inner and outer hair cells at the basal turn was 31.1% and 2.4%, respectively. In the ischemia/hypothermic group, the threshold returned to the pre-ischemic level within 30 min after reperfusion and remained stable thereafter. The mean loss of inner and outer hair cells on the seventh day was 0.1% and 0.2%, respectively. These results indicate that hypothermia can prevent inner ear damage, which otherwise occurs after transient ischemia of the cochlea.

Effect of calmodulin antagonists on the compound action potential of the cochlea

This study aimed to evaluate the effect of calmodulin antagonists on the threshold of the compound action potential (CAP) and the functional recovery of the cochlea after transient ischemia. When trifluoperazine and W-7 were administered to albino guinea pigs with perilymphatic perfusion, these drugs did not significantly affect the CAP thresholds. Transient cochlear ischemia of 30-min duration was obtained via a skull base approach. Although trifluoperazine significantly ameliorated the post-ischemic CAP threshold shifts 4 h after the onset of reperfusion, 1 to 50 μM W-7 did not affect the CAP threshold shifts. These results suggest that the action antagonizing calmodulin has no effect on the CAP threshold, while the role that calmodulin plays in cochlear ischemia-reperfusion injury still remains unclear.

Delayed Hearing Loss following Transient Cochlear Ischemia in the Gerbil

Delayed Hearing Loss following Transient Cochlear Ischemia in the Gerbil