Direct and indirect pathways for heterosynaptic interaction underlying developmental synapse elimination in the mouse cerebellum.

Developmental synapse elimination is crucial for shaping mature neural circuits. In the neonatal mouse cerebellum, Purkinje cells (PCs) receive excitatory synaptic inputs from multiple climbing fibers (CFs) and synapses from all but one CF are eliminated by around postnatal day 20. Heterosynaptic interaction between CFs and parallel fibers (PFs), the axons of cerebellar granule cells (GCs) forming excitatory synapses onto PCs and molecular layer interneurons (MLIs), is crucial for CF synapse elimination. However, mechanisms for this heterosynaptic interaction are largely unknown. Here we show that deletion of AMPA-type glutamate receptor functions in GCs impairs CF synapse elimination mediated by metabotropic glutamate receptor 1 (mGlu1) signaling in PCs. Furthermore, CF synapse elimination is impaired by deleting NMDA-type glutamate receptors from MLIs. We propose that PF activity is crucial for CF synapse elimination by directly activating mGlu1 in PCs and indirectly enhancing the inhibition of PCs through activating NMDA receptors in MLIs.

Tympanic membrane findings and Eustachian tube function after transtympanic plugging for the chronic patulous Eustachian tube.

OBJECTIVE: To evaluate Eustachian tube (ET) function after Kobayashi plug surgery based on the tympanic membrane (TM) findings and active opening (AO) of the ET assessed with sonotubometry. SUBJECTS AND METHODS: A retrospective survey of medical records identified 74 ears of 66 patients with patulous ET (PET) received transtympanic insertion of the Kobayashi plug. Excluding the six ears (6 patients) with abnormal preoperative TM, sixty-eight ears of 60 patients were found to have normal TM preoperatively. Among these 68 ears, there were 51 ears in which sonotubometry was performed both before and after surgery to evaluate whether the AO of the ET was positive or not. RESULTS: Out of the 68 ears with normal preoperative TM, 52 ears (76.5%) were judged successful (sum of complete relief and significant improvement). The postoperative TM was normal in 41 ears (60.3%), while 27 ears (39.7%) had abnormal TM findings postoperatively. The success rate was 75.6% (31/41) in ears with normal postoperative TM, while it was 77.8% (21/27) in ears with abnormal TM. Success in maintaining normal postoperative TM was found in 45.6% (31/68) of the total ears treated. Out of the 51 ears in which sonotubometry was performed both before and after surgery, AO was preoperatively positive in 88.2% of the ears (45/51), while it was positive in 64.7% (33/51) postoperatively. In thirty-four ears with normal TM postoperatively, AO was positive in 24 ears (70.6%), while it was positive in 9 out of 17 ears (52.9%) with abnormal postoperative TM. The success rate was 70.6% (36/51) for the 51 ears in which AO was assessed both pre- and postoperatively, and it was 66.7% (22/33) in ears with positive AO postoperatively, while it was 77.8% (14/18) in ears without AO postoperatively. The incidence of ears either having normal postoperative TM or positive AO postoperatively was 84.3% (43/51). Abnormal postoperative TM findings without effectiveness were found in 8.8% (6/68). CONCLUSION: The obstructive dysfunction of the ET is a calculated risk but did not occur in most ears after plugging with the Kobayashi plug. Therefore, routine insertion of the VT at the same time as the initial surgery is not recommended for PET cases that are adequately followed up.

Hearing Outcomes of Transmastoid Plugging for Superior Canal Dehiscence Syndrome by Underwater Endoscopic Surgery: With Special Reference to Transient Bone Conduction Increase in Early Postoperative Period.

OBJECTIVE: To investigate the influence on hearing of transmastoid plugging of the superior semicircular canal accompanied with membranous superior canal transection by underwater endoscopic ear surgery (UWEES) for the superior semicircular canal dehiscence syndrome. STUDY DESIGN: Retrospective case review. SETTING: Tertiary referral center. PATIENTS: Seven patients underwent plugging with membranous superior canal transection with UWEES from 2017 to 2019. INTERVENTION: Bone conduction (BC) thresholds (250, 500, 1000, 2000, 4000 Hz) were repetitively examined in early postoperative period. MAIN OUTCOME MEASURES: Subjective symptoms and pure-tone audiometry. RESULTS: Transient BC threshold increase was detected in all cases in early postoperative period and hearing levels were ameliorated in 1 to 2 months. The mean maximum BC threshold elevations (dB) during the early postoperative period (within 1 mo) and the postoperative stable hearing period (after 2 mo) were 18.6 and 2.9 at 250 Hz, 24.3 and 8.6 at 500 Hz, 26.4 and 8.6 at 1000 Hz, 28.6 and 7.1 at 2000 Hz, and 30.0 and 0.8 (except for scale-out cases) at 4000 Hz. respectively. The mean maximum BC thresholds in the early period were significantly elevated compared with those in the stable period at each frequency (p < 0.01). CONCLUSIONS: Hearing outcomes of transmastoid plugging with transection of the membranous superior canal by using UWEES were found favorable in a long-term follow-up. However, it caused transient reversible hearing loss in all cases. The BC increase in early postoperative period may not cause permanent hearing loss but improvement for the surgical technique may still be necessary.

Enlargement of Eustachian Tube Lumen in Patulous Eustachian Tube Patients Compared to that by Aging.

OBJECTIVE: To evaluate the Eustachian tube (ET) dimensions in patulous ET (PET) patients compared with that by aging using sitting 3D computed tomography (CT). STUDY DESIGN: Retrospective study. SETTING: Tertiary referral center. SUBJECTS AND METHODS: A retrospective survey of medical records in Sen-En Rifu Hospital identified 105 ears of 76 PET patients and 65 ears of 34 patients without ET dysfunction findings (non-PET). Subjects in both PET and non-PET groups were then divided into two age groups. Groups A and C defined as non-PET and PET subjects respectively, who were under the age of 60 years, while Groups B and D defined of non-PET and PET subjects respectively, who were 60 years and above. 3D CT (Accuitomo; Morita, Kyoto, Japan) was performed on all subjects in the sitting position. The ET lumen from the ET pharyngeal orifice to 15 mm was analyzed. RESULTS: No significant difference in the ET lumen near the pharyngeal orifice was found between Group A and C; however, there was a significant difference in the ET at points lateral to the pharyngeal orifice. The ET lumen was significantly larger at the site close to the pharyngeal orifice in Group B as compared to that of Group A. For the two groups of PET classified according to the size of the pharyngeal orifice and area close to the isthmus, age was only significantly different between two groups of pharyngeal orifice. On the contrary, sonotumometry and Ohta method were significantly different between the two groups of the area close to the isthmus. CONCLUSION: The lumen of the ET is enlarged in both PET and aging. However, the responsible site was found to be different. While the enlargement of the ET lumen in 60 years and above subjects without PET mainly occurred near the pharyngeal orifice of the ET, it was near the isthmus in under 60 years PET patients. Further study of possible clinical implications of these findings as well as treatment strategy are required.

Underwater Endoscopic Ear Surgery for Closure of Cholesteatomatous Labyrinthine Fistula With Preservation of Auditory Function.

OBJECTIVE: To analyze the outcomes of the underwater endoscopic ear surgery (UWEES) technique for closure of cholesteatomatous labyrinthine fistula (LF) with preservation of auditory function. STUDY DESIGN: Retrospective case review study. SETTING: Tertiary referral center. PATIENTS: A total of 12 patients with cholesteatomatous LF. INTERVENTION: Surgical method of closure using UWEES for cholesteatomatous LF to minimize inner ear damage. Artificial cerebrospinal fluid (CSF) was used as the perfusate, except for earlier cases when saline was employed. MAIN OUTCOME MEASURES: Comparison of bone conductance hearing level (BCHL) before and after surgery. A change of BCHL less than 10 dB was defined as successful preservation of bone conductance hearing. RESULTS: All cases of LF were treated successfully by closure using the UWEES technique. Seven cases were type I, one was type IIa, and four were type III according to the Milewski and Dornhoffer classification of LF. The average LF size was 3.1 mm (1-7 mm). Eleven patients were evaluated and their bone conductance hearing was well preserved in all of them (11/11). One patient was too young for preoperative evaluation of BCHL, but hearing preservation was verified 2 years later at the age of 6 years. Remarkably, none of the patients complained of vertigo, except for only a slight manifestation on postoperative day 1. CONCLUSION: The UWEES technique was effective for closure of cholesteatomatous LF with preservation of auditory function.

Patulous Eustachian Tube Patients With Respiratory Fluctuation of Tympanic Membrane in Both Sitting and Supine Positions: A Sign of Severity of Disease?

OBJECTIVE: The aim is to report four cases of patulous Eustachian tube (PET) patients with respiratory fluctuation of the tympanic membrane (TM) even in the supine position, and to examine the frequency and characteristics of such patients. PATIENTS: There were 195 ears (99 right ears and 96 left ears) from 146 cases (56 male and 90 female subjects aged 8-88, average 48.0 ±â€Š18.9 yrs) diagnosed with definite PET by diagnostic criteria proposed by Japan Otologic Society (JOS) between January 2017 and December 2019 at Sen-En Rifu Hospital. Patients who presented with respiratory fluctuation of the TM in both the sitting and supine positions were examined. MAIN OUTCOME MEASURES: Clinical patient records, the severity of subjective symptoms (PET handicap inventory-10 [PHI-10]), objective ET function tests (tubo-tympano-aerodynamic graphy [TTAG] and sonotubometry), and sitting three-dimensional computed tomography (3-D CT) were analyzed. RESULTS: Six ears (3.1%) of four cases (2.7%) exhibited respiratory fluctuation of the TM, even in the supine position. In these six ears (four cases), the PHI-10 score ranged from 16 to 36 with three ears exhibiting PHI-10 score equal to or exceeding 26 (in the category of severe handicap). Sitting CT indicated the findings of completely open ET in only two ears. All ears but one were managed by conservative treatment. CONCLUSION: Respiratory fluctuation of the TM in both the sitting and supine positions was observed in 2.7% of the definite PET patients. Surprisingly, such findings can be an indication of neither the subjective nor objective severity of the disease. Therefore, even for PET patients with such findings, surgery should not be immediately proposed, but rather, conservative management should be undertaken first.

Objective assessment of autophony during phonation in the diagnosis of patulous Eustachian tube patients.

OBJECTIVE: A system enabling the objective assessment of the transmission of voice sounds to the external auditory canal (EAC) during phonation has recently been revised. Our aim was to evaluate the effectiveness of this new system in the diagnosis of patulous Eustachian tube (PET) patients by comparing the results obtained using this method with those obtained from conventional objective tests to diagnose PET. METHODS: A prospective survey of medical records was included with definite PET, possible PET, and sensorineural hearing loss as control. The measurement system consists of a personal computer, an AD/DA converter (NI 6361, National Instruments), a probe microphone system for recording voice sound (ER-10C, Etymotic Research) and two microphones for measuring noise sound in the EAC (ER-10B+, Etymotic Research). Pronouncing the "Ni" sound for 5 s were recorded with these three microphones. The ratio of the maximum sound pressure of voice sound and noise sound in EAC (EAC/Voice) was simultaneously calculated, and results were displayed on a personal computer for diagnosing. RESULTS: Thirty-one patients of 42 ears with definite PET, 26 patients of 38 ears with possible PET, and 12 patients of 24 ears with sensorineural hearing loss as control were included. The EAC/Voice were 8.63 ± 5.43, 25.41 ± 32.63, and 25.87 ± 24.93 in the control, definite PET, and possible PET group respectively. The control group was significantly different from the definite PET (p < 0.05) and possible PET group (p < 0.05). ROC curve analysis confirmed 14.7 as the best diagnostic cut-off value of EAC/Voice (area under the curve=0.782, 95% CI 0.671-0.894). By adopting this cut-off point, 25 (56.8%) and 22 (61.1%) ears were determined as positive findings in the definite PET and possible PET group, respectively. There was no significant correlation between the positive findings judged by the current method and that of sonotubometry in the control (r = -0.63, p = 0.769), definite PET (r = 0.12, p = 0.451), and possible PET group (r = 0.12, p = 0.451). CONCLUSION: The current system is more useful in the objective assessment of autophony during phonation by calculating the ratio of voice sound and elicited noise sound transmitted in the EAC (EAC/Voice). This method seems promising because it is able to detect cases eluding conventionally used test methods such as sonotubometry performed without phonation, thereby increasing the accuracy of PET diagnoses.

Pulsatile tinnitus caused by pneumocephalus after Janneta surgery.

Pulsatile tinnitus of nonvascular origin is rare. We herein present a case of pulsatile tinnitus complicated with Jannetta surgery due to a communication created between the drilled mastoid cells and epidural space. She was successfully cured by otological surgery where the mastoid tip was packed with bone cement. A 68-year-old woman was referred to the previous hospital with complaints of right autophony, aural fullness, hyperacusis to her footsteps, and pulsatile tinnitus for the past three years. She had received Jannetta surgery for right hemifacial spasm seven years before. The computed tomography (CT) of the right temporal bone showed bony dehiscence between the mastoid cells and posterior cranial fossa. She underwent otological surgery to obliterate the tip of the mastoid cavity with artificial bone cement (BIOPEXⓇ) under general anesthesia. Her annoying aural symptoms were immediately abolished and she has been free from symptoms at ten months after surgery. It is critical to ensure the closure of any communication created between the middle ear and epidural space during surgeries in order to prevent the occurrence of pulsatile tinnitus.

The characteristic of patulous eustachian tube patients diagnosed by the JOS diagnostic criteria.

OBJECTIVE: The objective is to describe characteristics of patients diagnosed with patulous Eustachian tube (PET) using the Diagnostic Criteria proposed by Japan Otological Society, and to evaluate the efficiency of objective tests to determine patent Eustachian tube. STUDY DESIGN: Retrospective. SETTING: Tertiary referral center. SUBJECTS: A retrospective survey of medical records in Sen-En Rifu Hospital identified 78 ears of 56 patients with "Definite PET" diagnosed by the JOS Diagnostic Criteria between January 2017 and December 2017. METHOD: Initial diagnosis, aural symptoms (voice autophony, aural fullness and breathing autophony), tubal obstruction procedures (posture change and pharyngeal orifice obstruction) and objective findings (tympanic membrane movement, Tubo-Tympano-Aerodynamic Graphy (TTAG) and sonotubometry) were evaluated. In addition, sonotubometry with postural change (Ohta's method), sitting CT and a newly devised PHI-10 score were also examined. RESULTS: Voice autophony, aural fullness, and breathing autophony were observed in 93.6%, 87.2%, 78.2%, respectively. In 91% of the ears, PET symptoms improved by postural change from sitting to the lying / forward-bending position. Synchronous movement of the TM upon respiration was observed in 69.1% of the ears. Positive findings of TTAG were observed in 75.6% of ears. Positive findings of sonotubometry were found in 55.1% of ears. Sonotubometry with postural change (Ohta's method), when the cut-off value of over 10dB was used, was positive in 45.2% of ears. Newly devised PHI-10 score representing severity of subjective symptoms classifying patients into no handicap, mild handicap, moderate handicap and severe handicap were observed in 12.2%, 10.8%, 18.9% and 58.1% of ears, respectively. The evaluation of the extent of patency of the ET by sitting CT indicated completely open, closed-short, and closed-long, in 68.6%, 11.4% and 21.4% of ears, respectively. Compared to the closed group, the completely open group had a significantly higher frequency of positive breathing autophony, positive sonotubometry, and positive Ohta's method. CONCLUSION: The characteristics of main symptoms and the efficiency of various tests in PET diagnosis were analyzed based on data obtained from "Definite PET" patients diagnosed by the JOS Diagnostic Criteria. The greater the availability of tests to evaluate PET, the greater the opportunities to diagnose "Definite PET". In particular, tests measuring pressure transmission between the nasopharynx and middle ear, such as TM observation and TTAG, are more sensitive than sonotubometry measuring sound transmission.

Plug size selection protocol for the treatment of intractable patulous Eustachian tube with Kobayashi Plug.

Background: There have been no useful criteria for initial plug size selection protocol for the treatment of intractable patulous Eustachian tube (PET). Aims/objectives: To establish a method for appropriate plug size selection using tubal function test and subjective symptom severity in PET patients who were treated by Kobayashi Plug insertion. Material and methods: A retrospective survey of medical records identified 39 ears of 35 patients with PET who received insertion of the Kobayashi Plug and whose PET symptoms were thereafter controlled for at least 6 months after surgery. Method: The evaluation scale of PET handicap inventory-10 (PHI-10) was used to indicate PET subjective symptom severity. Tubal function tests (sonotubometry and tubo-tympano-aerodynamic-graphy: TTAG) were performed. Results: There was no correlation between the preoperative PHI 10 score and plug size (p = .157). There was a significant correlation between the preoperative sound attenuation from nostril to EAC measured by sonotubometry and plug size (p < .001). There was no correlation between the preoperative pressure transmission ratio estimated by TTAG and plug size (p = .271). Conclusions and Significance: Sonotubometry which evaluates sound attenuation from nostril to EAC can be a useful tool for selecting plug size.

Serotonin rebalances cortical tuning and behavior linked to autism symptoms in 15q11-13 CNV mice.

Serotonin is a critical modulator of cortical function, and its metabolism is defective in autism spectrum disorder (ASD) brain. How serotonin metabolism regulates cortical physiology and contributes to the pathological and behavioral symptoms of ASD remains unknown. We show that normal serotonin levels are essential for the maintenance of neocortical excitation/inhibition balance, correct sensory stimulus tuning, and social behavior. Conversely, low serotonin levels in 15q dup mice (a model for ASD with the human 15q11-13 duplication) result in impairment of the same phenotypes. Restoration of normal serotonin levels in 15q dup mice revealed the reversibility of a subset of ASD-related symptoms in the adult. These findings suggest that serotonin may have therapeutic potential for discrete ASD symptoms.

Spike timing-dependent selective strengthening of single climbing fibre inputs to Purkinje cells during cerebellar development.

Shaping functional neural circuits in developing brain involves activity-dependent refinement of early-formed redundant synapses. In the developing cerebellum, a one-to-one connection between a climbing fibre (CF) and a Purkinje cell (PC) is established by selective strengthening of a single CF followed by elimination of surplus CFs. Here we investigate developmental changes in CF-mediated responses in PCs by using in vivo whole-cell recordings and two-photon Ca(2+) imaging. We show that each neonatal PC receives temporally clustered inputs from multiple CFs and temporal integration of these inputs is required to induce burst spiking and Ca(2+) rise in PCs. Importantly, a single CF input closest to PC's spike output is selectively strengthened during postnatal development. This spike timing-dependent selective strengthening is much less prominent in PC-selective P/Q-type voltage-dependent Ca(2+) channel knockout mice. Thus, spike timing- and Ca(2+)-dependent plasticity appears to underlie the selection of a single 'winner' CF and the establishment of mature CF-PC connections.

The CB1 cannabinoid receptor is the major cannabinoid receptor at excitatory presynaptic sites in the hippocampus and cerebellum.

Endocannabinoids work as retrograde messengers and contribute to short-term and long-term modulation of synaptic transmission via presynaptic cannabinoid receptors. It is generally accepted that the CB1 cannabinoid receptor (CB1) mediates the effects of endocannabinoid in inhibitory synapses. For excitatory synapses, however, contributions of CB1, "CB3," and some other unidentified receptors have been suggested. In the present study we used electrophysiological and immunohistochemical techniques and examined the type(s) of cannabinoid receptor functioning at hippocampal and cerebellar excitatory synapses. Our electrophysiological data clearly demonstrate the predominant contribution of CB1. At hippocampal excitatory synapses on pyramidal neurons the cannabinoid-induced synaptic suppression was reversed by a CB1-specific antagonist, N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide (AM251), and was absent in CB1 knock-out mice. At climbing fiber (CF) and parallel fiber (PF) synapses on cerebellar Purkinje cells the cannabinoid-dependent suppression was absent in CB1 knock-out mice. The presence of CB1 at presynaptic terminals was confirmed by immunohistochemical experiments with specific antibodies against CB1. In immunoelectron microscopy the densities of CB1-positive signals in hippocampal excitatory terminals and cerebellar PF terminals were much lower than in inhibitory terminals but were clearly higher than the background. Along the long axis of PFs, the CB1 was localized at a much higher density on the perisynaptic membrane than on the extrasynaptic and synaptic regions. In contrast, CB1 density was low in CF terminals and was not significantly higher than the background. Despite the discrepancy between the electrophysiological and morphological data for CB1 expression on CFs, these results collectively indicate that CB1 is responsible for cannabinoid-dependent suppression of excitatory transmission in the hippocampus and cerebellum.

Adenosine A(1)-receptor-mediated tonic inhibition of glutamate release at rat hippocampal CA3-CA1 synapses is primarily due to inhibition of N-type Ca(2+) channels.

The voltage-gated Ca(2+) channels responsible for synaptic transmission at CA3-CA1 synapses are mainly P/Q- and N-types. It has been shown that tonic inhibition of transmission due to activation of adenosine A(1) receptors occurs at this synapse. We have recently developed a technique to monitor synaptically released glutamate which is based on synaptically induced glial depolarisation. Using this technique, we have examined the effects of different voltage-gated Ca(2+) channel blockers on glutamate release. Under conditions in which the adenosine A(1) receptor was not blocked, omega-AgaIVA (a P/Q-type voltage-gated Ca(2+) channel blocker) suppressed synaptically induced glial depolarisation to a greater extent than omega-CgTxGVIA (an N-type voltage-gated Ca(2+) channel blocker) did. In contrast, in the presence of an adenosine A(1) receptor antagonist, omega-AgaIVA was less effective at suppressing synaptically induced glial depolarisation than omega-CgTxGVIA. These results indicate that, in the absence of adenosine A(1) receptor-mediated tonic inhibition, the contribution of N-type is much greater than that of P-type, and that N-types are the primary target of tonic inhibition in normal conditions in which adenosine A(1) receptor-mediated tonic inhibition is present.

Glutamate release increases during mossy-CA3 LTP but not during Schaffer-CA1 LTP.

Abstract It is still a matter of dispute whether the expression of hippocampal long-term potentiation (LTP) is due to enhanced transmitter release or enhanced postsynaptic sensitivity. Recently we developed a novel method to monitor synaptically released glutamate. In this method, brain slice preparations are stained with a voltage-sensitive dye RH155 which preferentially stains glial cells, and synaptically induced glial depolarization (SIGD) are optically detected in the presence of the blockers for ionotropic glutamate receptors. We have previously shown that SIGD is due to uptake of synaptically released glutamate by glial glutamate transporters. Here we applied this method to examine change in glutamate release during hippocampal LTP. To examine mossy-CA3 LTP, stimulating electrodes were placed in dentate gyrus and tetanic stimulation was delivered in the presence of 50 micro m APV. The amplitude of SIGD after inducing LTP was significantly greater than that in control experiments in which tetanus was not delivered. The amplitude of SIGD after inducing LTP by a brief (3-5 min) application of 50 micro m forskolin was also significantly greater than that in control experiments. At the Schaffer-CA1 synapse, the change in the amplitude of SIGD during LTP induced either by 100 Hz tetanus LTP or 200 Hz tetanus was not significantly greater than that of control experiments. These results provide evidence for increased glutamate release from the presynaptic terminals as the expression mechanism for both tetanus-induced and forskolin-induced LTP at mossy-CA3 synapses, and evidence supporting a postsynaptic expression mechanism at Schaffer-CA1 synapses.

Optical bioimaging: from living tissue to a single molecule: optical detection of synaptically induced glutamate transporter activity in hippocampal slices.

Since voltage-sensitive dyes (VSDs) change their optical properties in response to membrane potential changes, non-invasive optical measurements of membrane excitation using VSDs have been applied to various types of excitable tissue. This article deals with the authors' recent work using this technique in addition to basic and technical information about VSD imaging. By applying this technology to hippocampal brain slices, we have found that astrocytes play indispensable roles in sequestering glutamate, maintenance of synaptic transmission, and neuronal survival. Finally, the potential of VSD imaging for drug development was discussed.