Biomedical subjects

J Keifer

Publications and source records attributed to J Keifer.

At least 19 recordsLinked to original sources

PubMed2004

Targeting of GLUR4-containing AMPA receptors to synaptic sites during in vitro classical conditioning.

The synaptic delivery of GluR4-containing AMPA receptors during in vitro classical conditioning of a neural correlate of an eyeblink response was examined by fluorescence imaging of punctate staining for glutamate receptor subunits and the presynaptic marker synaptophysin. There was a significant increase in GluR4-containing AMPA receptors to synaptic sites after conditioning as determined by colocalization of GluR4 subunit puncta with synaptophysin. Moreover, the trafficking of these receptor subunits requires NMDA receptor activation as it was blocked by D,L-2-amino-5-phosphonovaleric acid (AP-5). In contrast, colocalization of NR1 subunits with synaptophysin was stable regardless of whether the preparations had undergone conditioning or had been treated by AP-5. The enhanced colocalization of GluR4 and synaptophysin was accompanied by an increase in both the total number and size of puncta for both proteins, suggesting greater synthesis and aggregation during conditioning. Western blot analysis confirmed upregulation of synaptophysin and GluR4 following conditioning. These data support the hypothesis that GluR4-containing AMPA receptors are delivered to synaptic sites during conditioning. Further, they suggest coordinate presynaptic and postsynaptic modifications during in vitro classical conditioning.

2-Amino-5-phosphonovalerate↗

PubMed2004

Social stress and corticosterone regionally upregulate limbic N-methyl-D-aspartatereceptor (NR) subunit type NR(2A) and NR(2B) in the lizard Anolis carolinensis.

Social aggression in the lizard Anolis carolinensis produces dominant and subordinate relationships while elevating corticosterone levels and monoaminergic transmitter activity in hippocampus (medial and mediodorsal cortex). Adaptive social behavior for dominant and subordinate male A. carolinensis is learned during aggressive interaction and therefore was hypothesized to involve hippocampus and regulation of N-methyl-d-aspartate (NMDA) receptors. To test the effects of social stress and corticosterone on NMDA receptor subunits (NR), male lizards were either paired or given two injections of corticosterone 1 day apart. Paired males were allowed to form dominant-subordinate relationships and were killed 1 day later. Groups included isolated controls, dominant males, subordinate males and males injected with corticosterone. Brains were processed for glutamate receptor subunit immunohistochemistry and fluorescence was analyzed by image analysis for NR(2A) and NR(2B) in the small and large cell divisions of the medial and mediodorsal cortex. In the small granule cell division there were no significant differences in NR(2A) or NR(2B) immunoreactivity among all groups. In contrast, there was a significant upregulation of NR(2A) and NR(2B) subunits in the large pyramidal cell division in all three experimental groups as compared with controls. The results revealed significantly increased NR(2A) and NR(2B) subunits in behaving animals, whereas animals simply injected with corticosterone showed less of an effect, although they were significantly increased over control. Upregulation of NR(2) subunits occurs during stressful social interactions and is likely to be regulated in part by glucocorticoids. The data also suggest that learning social roles during stressful aggressive interactions may involve NMDA receptor-mediated mechanisms.

Aggression↗

PubMed2001

In vitro eye-blink classical conditioning is NMDA receptor dependent and involves redistribution of AMPA receptor subunit GluR4.

The classically conditioned vertebrate eye-blink response is a model in which to study neuronal mechanisms of learning and memory. A neural correlate of this response recorded in the abducens nerve can be conditioned entirely in vitro using an isolated brainstem-cerebellum preparation from the turtle by pairing trigeminal and auditory nerve stimulation. Here it is reported that conditioning requires that the paired stimuli occur within a narrow temporal window of <100 msec and that it is blocked by the NMDA receptor antagonist d,l-2-amino-5-phosphonovaleric acid. Moreover, there is a significant positive correlation between the levels of conditioning and greater immunoreactivity with the glutamate receptor 4 (GluR4) AMPA receptor subunit in the abducens motor nuclei, but not with NMDAR1 or GluR1. It is concluded that in vitro classical conditioning of an abducens nerve eye-blink response is generated by NMDA receptor-mediated mechanisms that may act to modify the AMPA receptor by increasing GluR4 subunits in auditory nerve synapses.

Abducens Nerve↗

PubMed2000

Immunocytochemical localization of glutamate receptor subunits in the brain stem and cerebellum of the turtle Chrysemys picta.

The regional distribution of ionotropic (AMPA and NMDA) and metabotropic (mGluR1alpha) glutamate receptor subunits was examined in the brain stem and cerebellum of the pond turtle, Chrysemys picta, by using immunocytochemistry and light microscopy. Subunit-specific antibodies that recognize NMDAR1, GluR1, GluR4, and mGluR1alpha were used to identify immunoreactive nuclei in the brain stem and cerebellum. Considerable immunoreactivity in the turtle brain stem and cerebellum was observed with regional differences occurring primarily in the intensity of staining with the antibodies. The red nucleus, lateral reticular nucleus and cerebellum labeled intensely for NMDAR1 and moderately for GluR1. The cerebellum also labeled strongly for mGluR1alpha. All of the cranial nerve nuclei labeled intensely for NMDAR1 and to varying degrees for GluR1, GluR4, and mGluR1alpha. Counterstaining revealed the presence of neuronal somata where there were no immunoreactive neurons in individual nuclei. This finding suggests that there are subpopulations of immunoreactive neurons within a given nucleus that bear different glutamate receptor subunit compositions. The results suggest that the glutamate receptor subunit distribution in the brain stem and cerebellum of turtles is similar to that reported for rats. Additionally, there is considerable colocalization of NMDA and AMPA receptors as revealed by light microscopy. These results have implications for the organization of neural circuits that control motor behavior in turtles, and, generally, for the function of brain stem and cerebellar neural circuits in vertebrates.

Abducens Nerve↗

PubMed2000

Comparison of cortically and subcortically controlled motor systems. II. Distribution of anterogradely labeled terminal boutons on intracellularly filled rubrospinal neurons in rat and turtle.

The present study examined the circuitry of the red nucleus of the Sprague-Dawley rat and the freshwater pond turtle, Chrysemys picta, by using intracellular cell filling combined with anterograde tract tracing. Although both species have a well-developed cerebellorubral system, they differ in that the red nucleus of rats receives direct input from the motor areas of the cerebral cortex, whereas turtles do not. However, a direct descending projection from the hypothalamus to the red nucleus of turtles has been described. The aim of this study was to elucidate the relative functional contributions of the cerebellum and descending inputs to motor signal generation in the red nucleus. The results show that the cellular distribution of cerebellar inputs on rubrospinal neurons is similar between the rat and turtle; these projections are observed on the soma and the proximal and distal dendrites. In contrast, the hypothalamic inputs in turtles occupy mainly the more distally located dendrites, similar to the position of the cortical inputs in rats. These findings suggest that, first, the cerebellar inputs are not spatially segregated from the cortical or hypothalamic inputs in rats or turtles, as far as can be determined by light microscopy. Second, there is specificity of input from the cortex in rats and hypothalamus in turtles onto the distal portions of the dendrites. The similarity in the organizational features of the mammalian and non-mammalian cerebellorubrospinal systems has implications for interpretations of the relative roles of the cerebellum and cerebral cortex in motor control.

Animals↗

PubMed1999

Morbidity and mortality for alternate-site anesthesia care.

The bibliography contains very few references from anesthesia literature and a preponderance of citations from emergency medicine, radiology and pediatrics journals. The reviewed publications include studies addressing the relation of drug type and dosages to morbidity and mortality in alternate site locations. These studies approach the issue of safety through retrospective analysis of techniques that are associated with acceptably low morbidity. The occurrence of adverse anesthetic events within formalized sedation programs is also reviewed. The low morbidity suggests that the trend of out of the operating room anesthetic and sedative use will continue. The ability of formalized sedation programs to establish and enforce standards of practice as well as to continually collect quality assurance data is a strong endorsement for their continuation.

Journal Article↗

PubMed1999

Context-sensitive half-time and anesthesia: how does theory match reality?

The pharmacokinetic model, which describes context-sensitive decrement times, has received validation through direct measurement. Further validation has come from the ability of targeted infusion schemes to obtain reliable and stable concentrations of a variety of intravenous drugs. A complete understanding of the concept will enable the appropriate selection of drugs and an appreciation of the strengths and limitations of current drug delivery systems.

Journal Article↗

PubMed1999

Properties of conditioned abducens nerve responses in a highly reduced in vitro brain stem preparation from the turtle.

Previous work suggested that the cerebellum and red nucleus are not necessary for the acquisition, extinction, and reacquistion of the in vitro classically conditioned abducens nerve response in the turtle. These findings are extended in the present study by obtaining conditioned responses (CRs) in preparations that received a partial ablation of the brain stem circuitry. In addition to removing all tissue rostral to and including the midbrain and cerebellum, a transection was made just caudal to the emergence of the IXth nerve. Such ablations result in a 4-mm-thick section of brain stem tissue that functionally eliminates the sustained component of the unconditioned response (UR) while leaving only a phasic component. We refer to this region of brain stem tissue caudal to the IXth nerve as the "caudal premotor blink region." Neural discharge was recorded from the abducens nerve following a single shock unconditioned stimulus (US) applied to the ipsilateral trigeminal nerve. When the US was paired with a conditioned stimulus (CS) applied to the posterior eighth, or auditory, nerve using a delay conditioning paradigm, a positive slope of CR acquisition was recorded in the abducens nerve, and CR extinction was recorded when the stimuli were alternated. Resumption of paired stimuli resulted in reacquisition. Quantitative analysis of the CRs in preparations in which the caudal premotor blink region had been removed and those with cerebellar/red nucleus lesions showed that both types of preparations had abnormally short latency CR onsets compared with preparations in which these regions were intact. Preparations with brain stem transections had significantly earlier CR offsets as more CRs terminated as short bursts when compared with intact or cerebellar lesioned preparations. These data suggest that a highly reduced in vitro brain stem preparation from the turtle can be classically conditioned. Furthermore, the caudal brain stem is not a site of acquisition in this reduced preparation, but it contributes to the sustained activity of both the UR and CR. Finally, the unusually short CR onset latencies following lesions to the cerebellum are not further exacerbated by removal of the caudal brain stem. These studies suggest that convergence of CS and US synaptic inputs onto the abducens nerve reflex circuitry may underlie acquisition in this reduced preparation, but that mechanisms that control learned CR timing arise from the cerebellorubral system.

Abducens Nerve↗

PubMed1998

Comparison of cortically and subcortically controlled motor systems: I. Morphology of intracellularly filled rubrospinal neurons in rat and turtle.

The rat and turtle differ markedly in major structural features of the corticocerebellorubrospinal circuitry. Although both species have a well-developed cerebellorubrospinal system, they differ in that a direct cerebral cortical input to the red nucleus is present only in the rat. The aim of the present study was to compare features of the soma and dendritic morphology of rubrospinal neurons that receive cortical input, as in rats, with those that do not, as in turtles. Intracellular Lucifer Yellow injections of neurons retrogradely labeled with Fast Blue in the rat or activity-dependent sulforhodamine-labeled neurons in the turtle were used to fill rubrospinal neurons in 150-200-microm-thick fixed sections. Images of filled neurons were imported into a computer to analyze quantitatively soma and dendritic morphology. The results show that rubrospinal soma size is slightly larger in the rat than in the turtle. However, analysis of the dendritic morphology, including total dendritic length, length of primary, secondary, and tertiary dendritic branches, and a Scholl analysis of dendritic branch intersections across concentric rings, demonstrated no significant differences between the two species. These findings suggest that the basic dendritic morphology of rubrospinal neurons may have been established early in phylogeny, preceding the evolution of cortical inputs. Alternatively, similar dendritic morphologies may have arisen due to the presence of other synapses in the turtle that occupy the sites of the cortical input in the rat. This comparative approach provides insights into the information processing capabilities of cortically versus subcortically controlled motor systems.

Animals↗

PubMed1998

Distribution of hypoglossal motor neurons innervating the prehensile tongue of the African pig-nosed frog, Hemisus marmoratum.

Using retrograde neuronal tracers, a study of the distribution of hypoglossal motor neurons innervating the tongue musculature was performed in the African pig-nosed frog, Hemisus marmoratum. This species is a radically divergent anuran amphibian with a prehensile tongue that can be aimed in three dimensions relative to the head. The results illustrate a unique rostrocaudal distribution of the ventrolateral hypoglossal nucleus and an unusually large number of motor neurons within this cell group. During the evolution of the long, prehensile tongue of Hemisus, the motor neurons innervating the tongue have greatly increased in number and have become more caudally distributed in the brainstem and spinal cord compared to other anurans. These observations have implications for understanding neuronal reconfiguring of motoneurons for novel morphologies requiring new muscle activation patterns.

Animals↗

PubMed1998

Evidence for a photosensitive region in the caudal mesencephalon of the turtle brain.

Using an in vitro brainstem-cerebellum preparation from the turtle Chrysemys picta, burst discharge was recorded from the abducens nerve when light was directed on the brainstem. This burst discharge likely represents a neural correlate of the eye-blink reflex. Increasing the intensity of the light stimulus reduced the response latency from a duration of many seconds to approximately 1-2 s. No response was recorded when the light source was covered. The response was present when infrared light was blocked, and it could only be produced when the light spectra contained wavelengths below approximately 550 nm. Lesion experiments reveal that the photosensitive area is located caudal to the trochlear nerve and rostral to the trigeminal nerve. Single-unit microelectrode recordings demonstrate that this region is tonically active in the dark and that activity is suppressed by light. Pharmacological results show that the light response is blocked by application of compounds that act as serotonergic antagonists, and that antagonists of noradrenergic receptors (alpha or beta) either have no effect or their actions are variable. Taken together, these data suggest that an isthmo-optic-like area in the turtle brain is photosensitive to direct light and activates neural circuits that control eye movements.

Adrenergic Antagonists↗

PubMed1998

Central trigeminal and posterior eighth nerve projections in the turtle Chrysemys picta studied in vitro.

Recent electrophysiological studies in the turtle Chrysemys picta have suggested that a neural correlate of the eye-blink reflex can be evoked in an in vitro brain-stem-cerebellum preparation by electrical rather than natural stimulation of the cranial nerves. Discharge recorded in the abducens nerve, which is similar to EMG recordings from extraocular muscles during eye retraction, is triggered by a brief electrical stimulus applied to the ipsilateral trigeminal nerve. Evidence also suggests that pairing a one-second electrical stimulus applied to the posterior eighth nerve immediately prior to a single shock to the trigeminal nerve results in classically conditioned abducens nerve discharge in response to the previously neutral eighth nerve stimulus. In view of these physiological findings, the aim of the present study was to examine the central projections of trigeminal and posterior eighth nerve inputs to elucidate the anatomical substrates that may underlie the in vitro eye-blink reflex and possible pathways involved in reflex conditioning. Neurobiotin (NB) or fluorescein dextran (FD) was pressure injected into the cut end of either the trigeminal or posterior eighth nerve of the in vitro brainstem-cerebellum. Following trigeminal nerve injections, both tracers showed label in the ipsilateral trigeminal nuclear complex. Direct projections to the ipsilateral principal and accessory abducens motor nuclei were observed, suggesting that the eye-blink reflex is monosynaptic. Trigeminal nerve axons were also observed to terminate in the ipsilateral cerebellar cortex. The results of the posterior eighth nerve injections showed axonal projections and terminals in the cochlear, vestibular and principal sensory trigeminal nuclei. Terminal label was also observed in the ipsilateral cerebellar cortex, deep cerebellar nuclei, and in the principal and accessory abducens motor nuclei. Results from the NB cases suggested transneuronal transport of this tracer substance, whereas the FD cases did not. The present findings suggest that convergence of trigeminal and posterior eighth nerve inputs occurs in the ipsilateral cerebellar cortex, the principal sensory trigeminal nucleus, and the principal and accessory abducens motor nuclei. These regions of convergence may therefore be considered as potential sites of synaptic modification during in vitro studies of the conditioned abducens nerve reflex.

Animals↗

PubMed1997

The cerebellum and red nucleus are not required for In vitro classical conditioning of the turtle abducens nerve response.

The role of the cerebellum during motor learning is a controversial issue. Many authors have suggested that the cerebellum and its connections with the red nucleus are essential for the acquisition of the conditioned eye blink reflex. Although there is little argument that the cerebellum is an important component to the generation of the conditioned response (CR), a number of studies have suggested that the cerebellum is not essential for conditioning. Using an in vitro model of the classically conditioned turtle abducens nerve response, we investigated the effect of cerebellar and red nucleus lesions on the acquisition, extinction, and reacquisition of CRs. Neural discharge was recorded from the abducens nerve after a single shock unconditioned stimulus (US) was applied to the ipsilateral trigeminal nerve. When the US was paired with a conditioned stimulus (CS) applied to the posterior eighth, or auditory, nerve, a positive slope of CR acquisition was recorded in the abducens nerve. After extinction stimuli in which the CS and US were alternated, the number of CRs decreased to near zero. When the CS and US were once again paired, reacquisition at a faster rate was recorded. The CRs showed unusual timing features compared with preparations in which the cerebellum was intact; they had significantly shorter latencies and showed burst-like responses. These data demonstrate that it is possible to classically condition this in vitro preparation in the absence of the cerebellum and red nucleus. However, the latencies of CRs were found to be dramatically altered in the cerebellar-lesioned preparations, suggesting that the cerebellum does play a role in the timing of the CR.

Abducens Nerve↗

PubMed1997

A hypothalamic projection to the turtle red nucleus: an anterograde and retrograde tracing study.

It is well known that the reptilian red nucleus lacks a descending motor cortical input to the red nucleus, but has a well-developed cerebellar input. The present study was undertaken to determine whether there is a descending rubral input that originates from the hypothalamus. Using an in vitro preparation from the turtle, injections of neurobiotin into the red nucleus resulted in retrograde labeling of neurons in the suprapeduncular nucleus of the hypothalamus. Injections of either neurobiotin or fluorescein dextran into the suprapeduncular nucleus resulted in anterograde labeling of axons and terminal boutons in the red nucleus. The majority of these terminations appeared to lie in the medial part of the red nucleus. These data have implications for the potential control of the somatic motor system of reptiles by limbic system inputs.

Animals↗

PubMed1996

Organization of face representation in the cingulate cortex of the rhesus monkey.

Neuroanatomical and electrophysiological methods were used to investigate the organization of face representation in the cingulate gyrus of four rhesus monkeys. Injections of fluorescent tracers placed into electrophysiologically defined sectors of the primary (M1) and supplementary (M2) motor cortices demonstrated that the rostral part of area 24c and the rostral part of area 23c send projections to the face representations of M1 and M2. Injections of biotinylated dextran amine involving the rostral part of area 24c and rostral part of area 23c demonstrated a direct projection from both areas to the facial nucleus of the pons. Our data suggest that areas 24c (M3) and 23c (M4) each contain a face representation which directly affects cortical as well as subcortical neural centers controlling facial activity.

Amidines↗

PubMed1996

Somatosensory and movement-related properties of red nucleus: a single unit study in the turtle.

Extracellular recordings were performed from turtle red nucleus neurons to examine their responsiveness to peripheral somatic stimulation and to study differences between rubral sensory and movement-related responses. In pentobarbital sodium-anesthetized or decerebrate turtles, red nucleus neurons could be divided into two categories based on their response characteristics. The first group, which included 87% of neurons studied, had low spontaneous rates of activity and responded with excitation to electrical stimulation of the spinal cord or the cerebellum, or during active movement of the contralateral limbs. Neurons in this category were likely to be rubrospinal cells. The remaining 13% of cells studied had higher rates of spontaneous discharge and were inhibited by electrical stimulation or during active movement. These cells might be rubral GABAergic interneurons. Single red nucleus neurons responded with excitation and/or inhibition to somatosensory stimulation. Unlike the motor fields, which were restricted to a single contralateral limb, red nucleus sensory receptive fields were wide and often bilaterally distributed. Rubral responsiveness to sensory stimulation was found to be significantly diminished during active limb movements, thereby suggesting that sensory inputs to the red nucleus are not used for the on-line modification of motor commands. Inactivation of the cerebellar cortex enhanced the sensory responsiveness of rubral neurons and expanded the size of red nucleus receptive fields. These results suggest that the red nucleus receives substantial sensory input, and that the cerebellar cortex can modify the flow of sensory information to the red nucleus.

Animals↗

PubMed1996

Effects of red nucleus inactivation on burst discharge in turtle cerebellum in vitro: evidence for positive feedback.

1. In behaving animals the red nucleus produces sustained action potential discharge during movements of the limbs. These bursts are thought to encode parameters of movement and thereby represent motor commands. Similar bursts can be recorded in the in vitro brain stem-cerebellum from the turtle. In this preparation, sustained discharge of red nucleus neurons was postulated to be generated by N-methyl-D-aspartate-mediated cellular mechanisms acting in combination with positive feedback in a recurrent cerebellorubral network. The present study was designed to test this positive feedback hypothesis. During recording of sustained discharge in the deep cerebellar nuclei and cortex, the red nucleus was reversibly inactivated by microinjection. The positive feedback hypothesis would be supported if activity in the cerebellum was attenuated by inactivation of the red nucleus. A nonrecurrent source of excitation would have to be postulated if cerebellar activity was unaffected. 2. Extracellular single-unit recordings were made from neurons in the deep cerebellar nuclei, cerebellar cortex, and vestibular nuclei. Burst discharges were evoked by brief electrical stimuli applied to the spinal cord that activated sensory structures. During inactivation of the red nucleus, sensory projections to the cerebellum that may evoke burst discharge were unaffected. Pressure microinjections of cobalt, lidocaine, gamma-aminobutyric acid (GABA), or 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) were used to reversibly inactivate the red nucleus. Saline injections were also tested. 3. Sustained discharge of all neurons recorded in the lateral cerebellar nucleus was greatly attenuated or blocked completely by injection of the pharmacological agents into the red nucleus. These effects were reversible. Of the recordings in the cerebellar cortex, 63% of these were blocked. All four compounds tested were effective blockers of the bursts, although the effects of GABA were less potent than the others. Saline injections into the red nucleus showed no effect. Burst discharges of single units recorded in either the medial cerebellar nucleus or the vestibular complex, which do not receive input from the red nucleus, showed no effect of red nucleus inactivation. 4. The results showed that sustained discharge in the cerebellum was significantly attenuated by inactivation of the red nucleus even though sensory input that may trigger the bursts was intact. These data support the hypothesis that sustained discharge in the cerebellorubral circuit is generated by a distributed neuronal network that uses positive feedback. The results have implications for mechanisms underlying normal brain function and some motor disorders.

Action Potentials↗

PubMed1995

In vitro classical conditioning of abducens nerve discharge in turtles.

In vitro classical conditioning of abducens nerve activity was performed using an isolated turtle brainstem-cerebellum preparation by direct stimulation of the cranial nerves. Using a delayed training procedure, the in vitro preparation was presented with paired stimuli consisting of a 1 sec train stimulus applied to the auditory nerve (CS), which immediately preceded a single shock US applied to the trigeminal nerve. Conditioned and unconditioned responses were recorded in the ipsilateral abducens nerve. Acquisition exhibited a positive slope of conditioned responding in 60% of the preparations. Application of unpaired stimuli consisting of CS-alone, alternate CS and US, or backward conditioning failed to result in conditioning, or resulted in extinction of CRs. Latencies of CR onset were timed such that they occurred midway through the CS. Activity-dependent uptake of the dye sulforhodamine was used to examine the spatial distribution of neurons labeled during conditioning. These data showed label in the cerebellum and red nucleus during conditioning whereas these regions failed to label during unconditioned responses. Furthermore, the principal abducens nucleus labeled heavily during conditioning. These findings suggest the feasibility of examining classical conditioning in a vertebrate in vitro brainstem-cerebellum preparation. It is postulated that the abducens nerve CR represents a behavioral correlate of a blink-related eye movement. Multiple sites of conditioning are hypothesized, including the cerebellorubral circuitry and brainstem pathways that activate the principal abducens nucleus.

Abducens Nerve↗