Robert V. Harrison

Loss of Fat4 disrupts PCP signaling and oriented cell division and leads to cystic kidney disease

Tissue organization in Drosophila is regulated by the core planar cell polarity (PCP) proteins Frizzled, Dishevelled, Prickle, Van Gogh and Flamingo. Core PCP proteins are conserved in mammals and function in mammalian tissue organization. Recent studies have identified another group of Drosophila PCP proteins, consisting of the protocadherins Fat and Dachsous (Ds) and the transmembrane protein Four-jointed (Fj). In Drosophila, Fat represses fj transcription, and Ds represses Fat activity in PCP. Here we show that Fat4 is an essential gene that has a key role in vertebrate PCP. Loss of Fat4 disrupts oriented cell divisions and tubule elongation during kidney development, leading to cystic kidney disease. Fat4 genetically interacts with the PCP genes Vangl2 and Fjx1 in cyst formation. In addition, Fat4 represses Fjx1 expression, indicating that Fat signaling is conserved. Together, these data suggest that Fat4 regulates vertebrate PCP and that loss of PCP signaling may underlie some cystic diseases in humans.

Reorganization of auditory cortex after neonatal high frequency cochlear hearing loss

Cochleotopic representation in cortex (AI) is extensively reorganized in cats having neonatal, bilateral high frequency cochlear hearing loss. Anterior areas of AI, normally devoted to high frequencies, contain neurons which are almost all tuned to one lower frequency. This frequency corresponds, at the level of the cochlea, to the border between normal and damaged haircell regions.

Toward a battery of behavioral and objective measures to achieve optimal cochlear implant stimulation levels in children.

Objectives: Children require audible and comfortable stimulation from their cochlear implants immediately after device activation. To accomplish this, a battery of objective measures may be needed that could include the electrically evoked stapedius reflex (ESR), compound action potential from the auditory nerve (ECAP), and/or auditory brain stem response (EABR). In the present study, the following specific research questions were asked: In children using cochlear implants, 1) Can the ECAP, EABR, and ESR be recorded at the time of cochlear implantation? 2) What is the feasibility of measuring the ECAP, EABR, and the ESR repeatedly without the use of sedation over the first year of implant use? 3) Do ECAP, EABR, and ESR thresholds or behavioral measures change over time? 4) What is the relation between ECAP, EABR, and ESR thresholds and behavioral measures of threshold and comfortably loud levels? Design: In 68 children, ECAP, EABR, and ESR responses as well as behavioral measures of stimulation threshold and maximum stimulation were recorded at regular intervals over the first year of implant use. In each child, responses were recorded to electrical pulses provided by three different electrodes along the implanted array. Visual inspections of the stapedius reflex (V-ESR) evoked by activation of the same three electrodes at the time of surgery were performed in an additional 20 children. Results: ECAP and EABR measures were obtained in more than 84% of electrodes tested and 89% of children tested both in the operating room at the time of implant surgery (OR) and after surgery in nonsedated children. ESRs were recorded by using immittance measures in more than 65% of electrodes tested and 67% of children tested by 3 mo of implant use, but this technique was less successful in the OR and during early stages of device use. V-ESRs and ECAP thresholds were higher in the OR than ESRs and ECAPs at postoperative recording times. EABR and ECAP thresholds did not significantly change over the first 6 and 12 mo of implant use, respectively, whereas ESR thresholds increased. Behavioral measures of threshold decreased over time, whereas maximum stimulation levels rose over time. Behavioral measures of threshold and loudness were highly correlated at all test times. ECAP, EABR, and behavioral measures were lower when evoked by an electrode at the apical end of the implanted array than by more basal electrodes. Behavioral thresholds could be predicted mainly by ECAP thresholds, whereas maximum stimulation levels could best be predicted by ESR thresholds; both were significantly affected by the age at implantation. Conclusions: A combination of nonbehavioral measures can aid in the determination of useful cochlear implant stimulation levels, particularly in young children and infants with limited auditory experience. These measures can be made in the operating room and can be repeated after surgery when needed. Correction factors to predict threshold stimulation levels should be based on ECAP thresholds or EABR thresholds if necessary. Correction factors should be made for at least one apical and mid-array electrode, should take into account the age of the child, and may have to be revised during the first year of implant use. Maximum stimulation levels may be best determined by using the ESR.

Activity-Dependent Developmental Plasticity of the Auditory Brain Stem in Children Who Use Cochlear Implants

Objectives 1) To determine if a period of early auditory deprivation influences neural activity patterns as revealed by human auditory brain stem potentials evoked by electrical stimulation from a cochlear implant. 2) To examine the potential for plasticity in the human auditory brain stem. Specifically, we asked if electrically evoked auditory potentials from the auditory nerve and brain stem in children show evidence of development as a result of implant use. 3) To assess whether a sensitive or critical period exists in auditory brain stem development. Specifically, is there an age of implantation after which there are no longer developmental changes in auditory brain stem activity as revealed by electrically evoked potentials? Design The electrically evoked compound potential of the auditory nerve (ECAP) and the electrically evoked auditory brain stem response (EABR) were recorded repeatedly during the first year of implant use in each of 50 children. The children all had pre- or peri-lingual onset of severe to profound sensorineural hearing loss and received their implants at ages ranging from 12 mo to 17 yr. All children received Nucleus cochlear implant devices. All children were in therapy and in school programs that emphasized listening and required the children to wear their implants consistently. Results Initial stimulation from the cochlear implant evoked clear responses from the auditory nerve and auditory brain stem in most children. There was no correlation between minimum latency, maximum amplitude, or slope of amplitude growth of initial responses with age at implantation for ECAP eN1, EABR eIII and eV components (p > 0.05). During the first year of implant use, minimum latency of these waves significantly decreased (p < 0.01, p < 0.0001, p < 0.0001, respectively). Neural conduction time, measured using the interwave latency of ECAP eN1-EABR eIII for lower brain stem and EABR eIII-eV for upper brain stem, decreased during the period of 6 to 12 mo of cochlear implant use (p < 0.01 (lower), p < 0.0001(upper)). The ECAP wave eN1 and the EABR wave eV showed significant increases in amplitude during time of implant use (p < 0.05 and p < 0.01, respectively). There were no correlations between the rate of interwave latency decrease and the rate of amplitude increases and the age at which children underwent implantation (p < 0.05). Conclusions Activity in the auditory pathways to the level of the midbrain can be evoked by acute stimulation from a cochlear implant. EABR measures are not influenced by any period of auditory deprivation. Auditory development proceeds once the implant is activated and involves improvements in neural conduction velocity and neural synchrony. Underlying mechanisms likely include improvements in synaptic efficacy and possibly increased myelination. The developmental plasticity that we have shown in the human auditory brain stem does not appear from EABR data to be limited by a critical period during childhood.

Cerebral Vascular Abnormalities in a Murine Model of Hereditary Hemorrhagic Telangiectasia

Background and Purpose— Hereditary hemorrhagic telangiectasia type 1 (HHT1) is an autosomal dominant vascular dysplasia caused by mutations in the endoglin gene and characterized by dilated vessels and arteriovenous malformations (AVMs). To understand the etiology of this disorder, we evaluated the cerebral vasculature of endoglin heterozygous (Eng+/−) mice, which represent the only animal model of HHT1. Methods— The cerebral vasculature of Eng+/− and Eng+/+ mice from C57BL/6 (B6) and 129/Ola (129) strains with a differential susceptibility to HHT1 was studied with corrosion casting. Casts were observed by scanning electron microscopy to detect malformations and evaluate arterial diameters and orientation of endothelial nuclei. Measurements were taken to assess relative constriction at arteriolar branching points and downstream relative dilatation. Results— Three of 10 Eng+/− mice demonstrated abnormal vascular findings including AVMs, while none of 15 Eng+/+ mice did. The incidence of relative constriction at arteriolar branching points was significantly less in both Eng+/− groups than in their Eng+/+ counterparts. The occurrence of relative dilatation was significantly greater in B6-Eng+/− than in B6-Eng+/+ mice. Endothelial nuclei were significantly rounder and deviated more from the direction of blood flow in Eng+/− than in Eng+/+ mice. Conclusions— Eng+/− mice showed significant structural alterations in cerebral blood vessels, indicating that the level of endoglin on endothelium is critical for maintenance of normal vasculature. Since endoglin haploinsufficiency is associated with HHT1, such changes in arteriolar structures might occur in HHT1 patients and predispose them to AVMs and their sequelae.

An Evoked Potential Study of the Developmental Time Course of the Auditory Nerve and Brainstem in Children Using Cochlear Implants

Central auditory responses to electrical stimulation from a cochlear implant were studied in 75 pre-lingually deafened children and 11 adults. Electrically evoked auditory brainstem response (EABR) latencies significantly decreased with duration of cochlear implant use and were not significantly affected by the age at implant activation. Significant decreases in early latency waves and interwaves occurred within the first 1–2 months of implant use, whereas longer term changes (6–12 months) were found for eV and eIII-eV, which measure activity in the more rostral brainstem. Comparisons to acoustically evoked auditory brainstem response (ABR) in children with normal hearing suggested shorter interwave EABR latencies, reflecting either distinct neural generators or increased neural synchrony, but similar rates of change in the later latency eV and eIII-eV with time in sound. In sum, normal-like development of the rostral auditory brainstem is promoted by cochlear implant use in children of a wide range of ages.

Measurement of cerebral blood volume in mouse brain regions using micro-computed tomography

Micro-computed tomography (micro-CT) is an X-ray imaging technique that can produce detailed 3D images of cerebral vasculature. This paper describes the development of a novel method for using micro-CT to measure cerebral blood volume (CBV) in the mouse brain. As an application of the methodology, we test the hypotheses that differences in CBV exist over anatomical brain regions and that high energy demanding primary sensory regions of the cortex have locally elevated CBV, which may reflect a vascular specialization. CBV was measured as the percentage of tissue space occupied by a radio-opaque silicon rubber that fills the vasculature. To ensure accuracy of the CBV measurements, several innovative refinements were made to standard micro-CT specimen preparation and analysis procedures. Key features of the described method are vascular perfusion under controlled pressure, registration of the micro-CT images to an MRI anatomical brain atlas and re-scaling of micro-CT intensities to CBV units with selectable exclusion of major vessels. Histological validation of the vascular perfusion showed that the average percentage of vessels filled was 93+/-3%. Comparison of thirteen brain regions in nine mice revealed significant differences in CBV between regions (p<0.0001) while cortical maps showed that primary visual and auditory areas have higher CBV than primary somatosensory areas.

Neonatal Cochlear Hearing Loss Results in Developmental Abnormalities of the Central Auditory Pathways

We have used animal models of long term neonatal cochlear hearing loss to study developmental plasticity of the central auditory pathways. Newborn chinchilla pups and feline kittens were treated with the ototoxic drug amikacin, so as to induce basal lesions in the cochlea. At maturity these animals were used in single unit electrophysiological mapping studies, in which the cochleotopic organization of primary auditory cortex (of the cat) and the inferior colliculus of the midbrain (in the chinchilla) were mapped. We have observed, both in the midbrain and auditory cortex, massive reorganization of frequency representation. Most striking were the presence of large monotonic regions (i.e. large areas in which all neurons have similar tuning properties). Cochlear lesions which involve inner hair cells clearly modify the normal development of cochleotopic representation in the midbrain and cortical regions. We suggest that similar abnormal patterns of frequency representation will exist in human subjects with long term neonatal hearing loss.

Carboplatin ototoxicity: an animal model

A new animal model of ototoxicity is presented using intravenous carboplatin in adult chinchillas. A range of physiological and morphological effects was produced using doses calculated from the recommended therapeutic range (200-400 mg/m2). Auditory thresholds to tone pips stimuli were monitored using brain stem evoked responses (ABR). Cochlear histopathology was studied by light microscopy (LM) and ultrastructural hair cell abnormalities investigated with scanning electronmicroscopy (SEM). Carboplatin in this animal model predominantly affected the inner hair cells. This may provide an important model for the study of selective loss of the main afferent input in the auditory system.

Three Distinct Auditory Areas of Cortex (AI, AII, and AAF) Defined by Optical Imaging of Intrinsic Signals

Using pure-tone sound stimulation, three separate auditory areas are revealed by optical imaging of intrinsic signals in the temporal cortex of the chinchilla (Chinchilla laniger). These areas correlate with primary auditory cortex (AI) and two secondary areas, AII and the anterior auditory field (AAF). We have distinguished AI on the basis of concurrent single-unit electrophysiological recording; neurons within the AI intrinsic signal region have short (<15 ms) onset-response latencies compared with neurons recorded in AII and the AAF. Within AI, AII, and AAF we have been able to define cochleotopic or tonotopic organization from the differences in intrinsic signal areas evoked by pure tones at octave-spaced frequencies from 500 Hz to 16 kHz. The maps in AI and AII are arranged orthogonal to each other.