GAD levels and muscimol binding in rat inferior colliculus following acoustic trauma.

Pharmacological studies of the inferior colliculus (IC) suggest that the inhibitory amino acid neurotransmitter gamma-aminobutyric acid (GABA) plays an important role in shaping responses to simple and complex acoustic stimuli. Several models of auditory dysfunction, including age-related hearing loss, tinnitus, and peripheral deafferentation, suggest an alteration of normal GABA neurotransmission in central auditory pathways. The present study attempts to further characterize noise-induced changes in GABA markers in the IC. Four groups (unexposed control, 0 h post-exposure, 42 h post-exposure, and 30 days post-exposure) of 3-month-old male Fischer 344 rats were exposed to a high intensity sound (12 kHz, 106 dB) for 10 h. Observed hair cell damage was primarily confined to the basal half of the cochlea. There was a significant decrease in glutamic acid decarboxylase (GAD(65)) immunoreactivity in the IC membrane fraction compared to controls (P<0.05) at 0 h (-41%) and 42 h (-28%) post-exposure, with complete recovery by 30 days post-exposure (P>0.98). Observed decreases in cytosolic levels of GAD(65) were not significant. Quantitative muscimol receptor binding revealed a significant increase (+20%) in IC 30 days after sound exposure (P<0.05). These data suggest that changes in GABA neurotransmission occur in the IC of animals exposed to intense sound. Additional studies are needed to determine whether these changes are a result of protective/compensatory mechanisms or merely peripheral differentiation, as well as whether these changes preserve or diminish central auditory system function.

Treating acute stroke patients with intravenous tPA. The OSF stroke network experience.

BACKGROUND AND PURPOSE: Since the FDA approved tissue plasminogen activator (tPA) in 1996 for acute ischemic stroke, few data have been obtained during the postmarketing phase, and applicability in rural hospitals does not exist. We attempt to examine the safety and outcome of intravenous tPA for acute ischemic stroke in the OSF Stroke Network. METHODS: Fifty-seven consecutive patients treated with tPA were examined from June 1996 through December 1998. Admission and discharge National Institute of Health Stroke Scales (NIHSS), modified Rankin Scales (MRS), and discharge disposition, as well as intracerebral hemorrhage and mortality rates, were compared. RESULTS: Of 20 network hospitals, 12 had the experience of administering tPA. No statistically significant differences in the variables recorded were observed for patients treated at the community hospitals versus those who received tPA at the tertiary medical center. In 35% of patients, tPA was initiated by an emergency room or primary care physician in consultation with an OSF neurologist. At discharge, 47% of the patients had minimal or no disability (MRS, 0 to 1), 44% had an NIHSS score of 0 or 1, 54% went home, 25% were transferred to in-patient rehabilitation, 12% went to a nursing or skilled-care facility, and 9% died. Intracerebral hemorrhage rate was 9%; 5% were symptomatic. CONCLUSIONS: tPA can be administered safely with good outcome at community and rural hospitals. The OSF Stroke Network can serve as a model to assist small community hospitals to set up stroke programs and deliver up-to-date, acute stroke therapies.

Age-related changes in GABA(A) receptor subunit composition and function in rat auditory system.

A decline in the ability to discriminate speech from noise due to age-related hearing loss (presbycusis) may reflect impaired auditory information processing within the central nervous system. Presbycusis may result, in part, from functional loss of the inhibitory neurotransmitter GABA. The present study assessed age-related changes of the GABA(A) receptor in the inferior colliculus of young-adult, middle-aged, and aged rats related to: (i) receptor subunit composition and (ii) receptor function. Western blotting was used to measure protein levels of selected GABA(A) receptor subunits in preparations obtained from the inferior colliculus of Fischer 344 and Fischer 344/Brown-Norway F1 hybrid rats. In both strains, the aged group exhibited significant increases in gamma1 subunit protein and a decrease in alpha1 subunit protein. To examine the functional consequence of this putative age-related subunit change, we measured the ability of exogenous GABA to flux/translocate chloride ions into microsac preparations derived from Fischer 344 inferior colliculus. GABA-mediated chloride influx was significantly increased in samples prepared from the inferior colliculus of aged animals. Together with previous studies, these results strongly suggest an age-related change in GABA(A) receptor composition. These changes may reflect a compensatory up-regulation of inhibitory function in the face of significant loss of presynaptic GABA release. These findings provide one example of plastic neurotransmitter receptor changes which can occur during the ageing process.

Morphological deficits in noradrenergic neurons in GEPR-9s stem from abnormalities in both the locus coeruleus and its target tissues.

The epileptic condition of the genetically epilepsy-prone rat (GEPR) appears to be caused partially by deficiencies in the locus coeruleus (LC) innervation of the superior colliculus (SC). Previous studies provide quantitative documentation of noradrenergic morphological deficits in the moderately epileptic GEPR-3. The present findings extend these studies by applying cell culture methodology to assessments of the severely epileptic GEPR-9. Our data show that total neurite length, the number of neurite branch points per cell, the cross-sectional area of cell bodies, and the cell perimeter are deficient in noradrenergic neurons in LC + SC cocultures derived exclusively from GEPR-9s compared to analogous cocultures obtained solely from nonepileptic control rats. Partial restoration of LC neuron morphology toward normal occurs when the GEPR-9 SC component of the coculture is replaced with nonepileptic control SC. Finally, when the GEPR-9 SC is cocultured with the control LC, a partial morphological deficit occurs in the otherwise normal noradrenergic neurons. However, the magnitude of this deficit is less than that observed in noradrenergic neurons of the GEPR-9 LC cocultured with the control SC. These data support the hypothesis that the developmental deficiencies of noradrenergic neurons of the GEPR-9 are derived from two sources, the LC and its target tissue, in this case, the SC. Also, intrinsic abnormalities of the LC appear to make a more pronounced contribution to the noradrenergic deficits than do those which reside in the SC.

Age-related changes in [3H]strychnine binding in the vestibular nuclei of rats.

Glycine plays an important role as a neurotransmitter in the four vestibular nuclei (VN). The objective of this study was to determine if the levels of glycine-receptor binding in the VN change as a function of age. Quantitative receptor autoradiography was performed on brainstem sections from three age groups (3, 18 and 26 months) of Fischer 344 rats to assess binding in the VN. Glycine receptors were localized using [3H]strychnine binding. Strychnine binding declined monotonically with increasing age, such that the level of strychnine binding in each of the VN in the 28-month-old animals was approximately one-half that in the 3-month-olds. The age-related decrease in levels of strychnine binding suggest altered glycinergic function in the VN, which may in turn contribute to disturbances in equilibrium observed in the elderly.

Carbamazepine-induced release of serotonin from rat hippocampus in vitro.

PURPOSE: Carbamazepine is one of several antiepileptic drugs (AEDs) that release the inhibitory neurotransmitter serotonin as part of their pharmacodynamic action on brain neurons. We undertook this study to investigate the cellular processes by which carbamazepine (CBZ) releases serotonin from brain tissue. METHODS: Tissue slices were prepared from hippocampi of Sprague-Dawley rats. These hippocampal slices were preincubated in vitro in a buffer so that neurons within the slice would take up tritium-labeled serotonin. Subsequently the slices were superfused with buffer containing CBZ or other chemicals (or both) that increase the overflow of serotonin radioactivity. RESULTS: Carbamazepine produced a concentration-dependent (50, 125, 250, or 500 microM) increase in basal overflow of serotonin radioactivity from superfused rat hippocampal slices in vitro. In contrast, these concentrations did not alter potassium-stimulated release, suggesting that the CBZ-induced release does not depend on depolarization or exocytosis. Blockade of the neuronal membrane serotonin transporter by fluoxetine (1 microM) or citalopram (2 microM) did not alter overflow of serotonin radioactivity produced by 250 microM CBZ. p-chloramphetamine (10 microM) produced a substantial increase in overflow of serotonin radioactivity, and this effect appears to be antagonized by 250 microM CBZ. Uptake of [3H]-labeled serotonin into hippocampal synaptosomes was inhibited by CBZ with a median inhibitory concentration (IC50) of 511+/-33 microM and a Hill coefficient of 0.87+/-0.11, suggesting competitive inhibition of uptake by CBZ. CONCLUSIONS: We conclude that CBZ (a) releases serotonin from hippocampal slices independent of exocytosis and by a mechanism not involving the neuronal membrane serotonin transporter, and (b) at high enough concentration, blocks the neuronal serotonin transporter.

Glycine immunoreactivity and receptor binding in the cochlear nucleus of C57BL/6J and CBA/CaJ mice: effects of cochlear impairment and aging.

Glycinergic neurons in the cochlear nucleus (CN) of C57BL/6J (C57) and CBA/CaJ (CBA) mice were studied by using immunocytochemical and receptor-binding techniques. Adult C57 mice exhibit progressive cochlear pathology as they age, whereas aging CBA mice retain good hearing. In the CN of old C57 mice (18 months) with severe hearing loss, the number of glycine-immunoreactive neurons decreased significantly. The number (Bmax) of strychnine-sensitive glycine receptors (GlyR) decreased significantly in the dorsal CN of old C57 mice. Significant effects were not observed in the CN of middle-aged C57 mice (with less-severe hearing loss) or in very old CBA mice (which do not exhibit severe hearing loss). The data suggest that the combination of severe hearing loss and old age results in deficits in one or more inhibitory glycinergic circuits in the CN.

Alterations of GABAA receptor subunit mRNA levels in the aging Fischer 344 rat inferior colliculus.

The inhibitory neurotransmitter, gamma-aminobutyric acid (GABA) is critically involved in shaping neuronal responses to simple and complex acoustic stimuli in the central auditory structure, the inferior colliculus (IC). Studies in rat and human suggest that age-related changes in markers for GABA neurotransmission occur in the IC. In particular, these changes include findings indicative of an age-related increase in the efficacy/potency of GABA to inhibit ligand binding at the GABAA receptor picrotoxin site in the Fischer (F344) rat IC. Such changes in GABAA receptor modulation suggest the potential for an alteration in GABAA receptor subunit composition in the old rat IC. To test this idea, the present study used in situ hybridization to quantify age-related changes in GABAA receptor subunit mRNA levels in the three major subdivisions of the IC in the F344 rat: dorsal cortex (DCIC), external cortex (ECIC), and the central nucleus (CIC). In support of earlier findings of an age-related change in GABAA receptor modulation, the present study found: (1) GABAA receptor subunit mRNA levels were significantly altered in the IC of old rats, and (2) age-related changes in subunit levels appeared to be regionally selective and subunit specific. A highly significant increase in the level of the gamma 1 subunit mRNA was observed with little change in the levels of the alpha 1, beta 2, and gamma 2 subunit mRNAs. A nonstatistically significant increase in alpha 2 subunit mRNA was also observed. This observed increase in alpha 2 subunit mRNA could be important since previous expression studies have shown that the alpha 2 and gamma 1 subunits coassemble and are incorporated into GABAA receptors which appear to be more sensitive to GABA. If the observed changes in subunit mRNA levels with age correlate well with enhanced GABAA receptor function in the IC of old rats, this, in turn, may represent a compensatory mechanism in response to presynaptic GABAergic changes.

GABAA receptor binding in the aging rat inferior colliculus.

The inhibitory neurotransmitter GABA has been shown to be critically involved in shaping neuronal responses to simple and complex acoustic stimuli in the inferior colliculus. Studies in the rat and human inferior colliculus have suggested significant changes in functions related to GABA neurotransmission occur in the aged. These changes include significant decreases in GABA content, GABA release, GABA neurons, glutamate decarboxylase enzymatic activity, and GABAB receptor binding. Such changes within the inferior colliculus may affect the ability of elderly listeners to process complex acoustic signals, particularly in the presence of background noise. The present study was designed to examine the regional distribution and effects of aging on GABAA receptor binding sites in the Fischer 344 rat inferior colliculus using in vitro quantitative receptor autoradiography. [3H]GABA binding to GABAA receptors was significantly reduced in the inferior colliculus of young adult (3 months) and aged (18-26 months) rats when compared to 2-month animals. However, no significant changes were observed after 3 months of age. Single concentrations of tritiated GABAA receptor ligands (muscimol, t-butylbicycloorthobenzoate, and flunitrazepam) revealed no significant age-related changes in receptor binding in the inferior colliculus between 3 and 26 months of age. To characterize further the pharmacology of the GABAA receptor in the inferior colliculus, GABA modulation of the picrotoxin binding site was examined using [3H]t-butylbicycloorthobenzoate. When increasing concentrations of GABA were added to the incubation buffer, a significant decrease in binding was observed in the inferior colliculus of rats in each age group. In aged rats, the dose-response curve was shifted to the left, indicating an increase in the potency of GABA to inhibit [3H]t-butylbicycloorthobenzoate binding. Although no changes in GABAA receptor binding were detected in the inferior colliculus after 3 months of age, a significant alteration in interaction between the GABA and picrotoxin binding sites was observed in the inferior colliculus of aged rats when compared to 3-month-old young adults. This difference appears to reflect an increased sensitivity of the receptor to GABA modulation in aged rats and, thus, may serve as a compensatory mechanism to enhance GABAA receptor function in response to a presynaptic loss of inhibition.

Age-related reduction of [3H]strychnine binding sites in the cochlear nucleus of the Fischer 344 rat.

The present study used quantitative receptor autoradiography to examine the effects of aging on the binding profile of the strychnine-sensitive glycine receptor in the Fischer 344 rat. Glycine receptor binding sites were localized using [3H]strychnine in two principal subdivisions of the cochlear nucleus; the dorsal and anteroventral cochlear nucleus. These central auditory brainstem structures are known to receive extensive glycinergic inputs. In young rats, single concentrations of [3H]strychnine showed significantly higher binding levels in the dorsal cochlear nucleus than the anteroventral cochlear nucleus (+38%, P < 0.001). Little binding was detected in regions of the posteroventral cochlear nucleus, and no specific binding was apparent in the cerebellum. Saturation analysis in the dorsal cochlear nucleus revealed an affinity constant (Kd) of 16.9 nM and a maximum number of binding sites of 850 fmol/mg protein. A significant age-related decrease in [3H]strychnine (8 nM) binding was observed in the anteroventral cochlear nucleus (-37%, P = 0.003) and dorsal cochlear nucleus (-23%, P = 0.034) of 26-month-old rats compared with three-month-old rats. Saturation analysis indicated that the observed decrease in binding was due to a decrease in the total number of binding sites with no significant change in affinity. In the dorsal cochlear nucleus, the number of binding sites was reduced (-26%) in 26-month-old rats compared with three-month-old adults (P = 0.011). Kd was decreased (-22%) in 26-month-old rats when compared with young adults, but this decrease was not statistically significant (P = 0.377).(ABSTRACT TRUNCATED AT 250 WORDS)

Central auditory aging: GABA changes in the inferior colliculus.

Age-related hearing loss (presbycusis) is a complex state that reflects pathologic changes along the entire auditory neuraxis. Loss of speech understanding, decreased ability to localize sounds, and a decreased ability to detect and extract signals in noise are characteristic problems encountered by the elderly. Central (neural) presbycusis frequently results in a dramatic loss in speech understanding without a parallel change in pure-tone thresholds. In spite of evidence that suggests these deficits cannot be fully explained by peripheral changes alone, few studies have examined the neurochemical basis of central auditory dysfunction in aging. Age-related alterations in neural circuits involved in the processing of acoustic information could reflect changes in the synthesis, degradation, uptake, release, and receptor sensitivity of neurotransmitters, perhaps secondary to cell loss and/or progressive deafferentation. A series of studies designed to test this hypothesis has examined aging in the central auditory system of the F344 rate. Age-related changes associated with GABA neurotransmitter function in an important auditory midbrain structure, the inferior colliculus, have been investigated. These studies found: (1) decreased numbers of GABA immunoreactive neurons; (2) decreased basal levels (concentrations) of GABA; (3) decreased GABA release; (4) decreased glutamic acid decarboxylase activity; (5) decreased GABAB receptor binding; (6) decreased numbers of presynaptic terminals; and (7) subtle GABAA receptor binding changes. Collectively, these age-related changes suggest altered GABA neurotransmitter function in the IC. Identification of specific neurotransmitter changes in structures important in speech processing could eventually lead to the development of pharmacotherapy for selective types of age-related hearing loss.

Age-related decrease in GABAB receptor binding in the Fischer 344 rat inferior colliculus.

Quantitative receptor autoradiography was used to assess GABAB receptor binding in three primary subdivisions of the inferior colliculus (IC): dorsal cortex (DCIC), external cortex (ECIC), and the central nucleus (CIC) of 3-, 18-20-, and 26-month-old Fischer 344 rats. GABAB binding sites were localized using [3H]GABA in the presence of a saturating concentration of isoguvacine, a selective GABAA receptor agonist, to displace [3H]GABA bound to GABAA receptor sites. In the three IC subdivisions examined, GABAB receptor binding was significantly reduced in 26-month-old rats when compared to 3-month-old rats (DCIC, -44%; ECIC, -36%; CIC, -32%; p < 0.05). For comparison, GABAB binding was determined in the portion of cerebellum located in the recess of the IC. In the molecular layer of this region, there was no statistically significant differences in receptor binding between 3, 18-20-, and 26-month-old rats. In addition, there was not a significant age-related change in the cross-sectional area of the IC. These findings provide additional evidence to support the existence of selective age-related changes in GABA neurotransmitter function in the rat IC.

Age-related changes in brainstem auditory neurotransmitters: measures of GABA and acetylcholine function.

This study was designed to determine if there are age-related alterations in the bio-synthetic enzyme glutamic acid decarboxylase (GAD), the degradative enzyme GABA-transaminase (GABA-T), and the uptake system for GABA in the central nucleus of the inferior colliculus (CIC), the cochlear nucleus (CN), and/or nuclei of the lateral lemniscus (NLL) of Fischer-344 rats. For purposes of comparison, the cholinergic neuronal system was studied in parallel in young adult (3-7 months), mature (15-17 months) and aged (24-26 months) rats. In young adults GAD activity was highest in the CIC (219 nmol/mg protein/h; N = 5), intermediate in NLL (82 nmol/mg protein/h), and lowest in CN (34 nmol/mg protein/h). Choline acetyltransferase (ChAT) activity was highest in NLL and CN, and approximately 35-40% lower in CIC. A more uniform pattern was observed with GABA-T activity. Reductions in GAD activity were seen in the CIC of mature (-31%) and aged (-30%) rats that were not graded with age when compared to young adult, P < 0.05 (N = 5). This effect was regionally selective, since the CN did not show any loss of GAD or ChAT activity. The neurotransmitter selectivity of this deficit in CIC is supported by the non-parallel changes in ChAT activity (-22%, aged vs. mature, P < 0.05) that occurred after the changes in GAD activity. In contrast to the loss of GABAergic biosynthetic capacity in aged CIC, high affinity uptake processes (Kd and Vmax) for 14C-GABA and 3H-D-aspartate were not significantly altered (P > 0.05). Similar to the CIC, the NLL showed remarkable age-related deficits, but these deficits were more substantial for the cholinergic system (ChAT activity: -56% aged vs. young adult, P < 0.05; GAD activity: -35% aged vs. mature). None of the areas examined showed a significant loss of GABA-T activity with aging. These data suggest: 1) Age-related loss of GABA-mediated inhibition in the CIC of Fischer-344 rats is not attributable to changes in uptake or degradation of GABA, but may be related loss of biosynthetic capacity (i.e. activity or quantity) of the GAD present; 2) processing centers of the central auditory pathway (i.e. CIC and NLL), but not necessarily primary (i.e. CN) integrative nuclei, demonstrate selective, age-related neurochemical deficits; and 3) age-related neurochemical changes in central auditory structures may contribute substantially to the abnormal perception of signals in noise and loss of speech discrimination observed in neural presbycusis.