Safety and efficacy of leflunomide in the treatment of lupus nephritis refractory or intolerant to traditional immunosuppressive therapy: an open label trial.
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Adult↗
Biomedical subjects
W-C Li
Publications and source records attributed to W-C Li.
Glutamate and acetylcholine corelease at developing synapses.
Most neurons release a single fast-acting low-molecular-weight transmitter at synapses to activate and open postsynaptic ion channels. We challenge this principle with evidence for corelease of the two major excitatory transmitters, glutamate and acetylcholine (ACh), from single identified neurons in the developing frog tadpole spinal cord. Whole-cell patch electrodes were used to record from single spinal neurons. When action potentials and inhibition were blocked, spontaneous miniature excitatory postsynaptic currents (mEPSCs) were recorded. These were fully blocked only by joint application of glutamate [alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) and N-methyl-D-aspartate receptor (NMDAR)] and nicotinic ACh receptor (nAChR) antagonists. Fast nAChR and slow NMDAR mEPSCs were isolated pharmacologically. We then show that some mEPSCs have both the fast nAChR rise and slow NMDAR decay and conclude that some individual synaptic vesicles corelease glutamate and ACh. Whole-cell recordings from pairs of neurons were then used to identify the spinal interneurons coreleasing the two excitatory transmitters. One anatomical class of interneuron with descending axons was found to excite other spinal neurons by activating nAChR, AMPAR, and NMDAR simultaneously at its synapses. Although Jonas, Bischofberger, and Sandkuhler [Jonas, P., Bischofberger, J. & Sandkuhler, J. (1998) Science 281, 419-424] showed that the inhibitory transmitters GABA and glycine can be coreleased at spinal synapses, the Xenopus tadpole provides a case where the two main CNS excitatory transmitters are released from single vesicles, and where the presynaptic neuron coreleasing two transmitters has been identified.
Acetylcholine↗
Primitive roles for inhibitory interneurons in developing frog spinal cord.
Understanding the neuronal networks in the mammal spinal cord is hampered by the diversity of neurons and their connections. The simpler networks in developing lower vertebrates may offer insights into basic organization. To investigate the function of spinal inhibitory interneurons in Xenopus tadpoles, paired whole-cell recordings were used. We show directly that one class of interneuron, with distinctive anatomy, produces glycinergic, negative feedback inhibition that can limit firing in motoneurons and interneurons of the central pattern generator during swimming. These same neurons also produce inhibitory gating of sensory pathways during swimming. This discovery raises the possibility that some classes of interneuron, with distinct functions later in development, may differentiate from an earlier class in which these functions are shared. Preliminary evidence suggests that these inhibitory interneurons express the transcription factor engrailed, supporting a probable homology with interneurons in developing zebrafish that also express engrailed and have very similar anatomy and functions.
Action Potentials↗
Dorsal spinal interneurons forming a primitive, cutaneous sensory pathway.
In mammals, sensory projection pathways are provided by just three classes of spinal interneuron that develop from the roof-plate. We asked whether similar sensory projection interneurons are present primitively in a developing lower vertebrate where function can be more readily studied. Using an immobilized Xenopus tadpole spinal cord preparation, we define the properties and connections of spinal sensory projection interneurons using whole cell patch recordings from single neurons or pairs, identified by dye filling. Dorsolateral interneurons lie in the tadpole equivalent of the spinal dorsal horn, and have dorsally located dendrites and an ipsilateral ascending axon that projects into the midbrain. These neurons receive direct, mainly AMPA receptor (AMPAR)-mediated, excitation from skin touch sensory neurons. They in turn produce AMPAR and N-methyl-d-aspartate receptor (NMDAR)-mediated excitation of spinal locomotor pattern generator neurons rostrally on the same side of the cord. During swimming, they receive glycinergic modulatory inhibition from ascending interneurons in the spinal locomotor central pattern generator. We conclude that spinal dorsolateral interneurons are a primitive class of excitatory sensory projection neurons activated by ipsilateral cutaneous afferents and carrying excitation ipsilaterally and rostrally as far as the midbrain to initiate or accelerate swimming.
Action Potentials↗
A direct comparison of whole cell patch and sharp electrodes by simultaneous recording from single spinal neurons in frog tadpoles.
High-impedance, sharp intracellular electrodes were compared with whole cell patch electrodes by recording from single spinal neurons in immobilized frog tadpoles. A range of neuron properties were examined using sharp or patch test electrodes while making simultaneous recordings with a second control patch electrode. Overall, test patch electrodes did not significantly alter the activity recorded by the control electrode, and recordings from the two electrodes were essentially identical. In contrast, sharp electrode recordings differed from initial control patch recordings. In some cases, differences were due to real changes in neuron properties: the resting membrane potential became less negative and the neuron input resistance (R(i)) fell; this fall was larger for neurons with a higher R(i). In other cases, the control patch electrode revealed that differences were due to the recording properties of the sharp electrode: tip potentials were larger and more variable; resting potentials appeared to be more negative; and spike amplitude was attenuated. However, sharp electrode penetration did not, in most cases, significantly alter the pattern of neuron firing in response to injected current or the normal pattern of activity following sensory stimulation or during fictive swimming. We conclude that sharp electrodes introduce a significant leak to the membrane of tadpole spinal neurons compared with patch electrodes but that this does not change the fundamental firing characteristics or activity of the neurons.
Action Potentials↗
Brainstem control of activity and responsiveness in resting frog tadpoles: tonic inhibition.
The hatchling Xenopus laevis tadpole was used to study the brain neurons controlling responsiveness. Tadpoles have reduced motor activity and responsiveness when they hang at rest, attached by cement gland mucus. Afferent input from cement gland mechanosensory neurons has both a phasic role in stopping swimming and a tonic role in reducing responsiveness while tadpoles hang attached. Both these roles depend on GABA(A)-mediated inhibition. We provide evidence supporting the hypothesis that long-term reduced responsiveness in attached tadpoles results from tonic activity in the reticulospinal GABAergic pathway mediating the stopping response. Two groups of putative stopping pathway interneurons were recorded in the caudal and rostral hindbrain of immobilised tadpoles. Both groups showed a sustained increase in activity during simulated attachment. This attached activity was irregular and unstructured. We consider whether low-level firing in cement gland afferents (at approximately 1 Hz) during simulated attachment is sufficient to explain the low-level firing (at approximately 0.5 Hz) in reticulospinal neurons. We then ask if a small population of these neurons (approximately 20) could produce sufficient inhibition of spinal neurons to reduce the whole tadpole's responsiveness. We conclude that for most of their 1st day of life GABAergic brainstem neurons could produce inhibition continuously while the tadpole is at rest.
Action Potentials↗
Spinal inhibitory neurons that modulate cutaneous sensory pathways during locomotion in a simple vertebrate.
During locomotion, reflex responses to sensory stimulation are usually modulated and may even be reversed. This is thought to be the result of phased inhibition, but the neurons responsible are usually not known. When the hatchling Xenopus tadpole swims, responses to cutaneous stimulation are modulated. This occurs because sensory pathway interneurons receive rhythmic glycinergic inhibition broadly in phase with the motor discharge on the same side of the trunk. We now describe a new whole-cell recording preparation of the Xenopus tadpole CNS. This has been used with neurobiotin injection to define the passive and firing properties of spinal ascending interneurons and their detailed anatomy. Paired recordings show that they make direct, glycinergic synapses onto spinal sensory pathway interneurons, and the site of contact can be seen anatomically. During swimming, ascending interneurons fire rhythmically. Analysis shows that their firing is more variable and not as reliable as other interneurons, but the temporal pattern of their impulse activity is suitable to produce the main peak of gating inhibition in sensory pathway interneurons. Ascending interneurons are not excited at short latency after skin stimulation but are strongly active after repetitive skin stimulation, which evokes vigorous and slower struggling movements. We conclude that ascending interneurons are a major class of modulatory neurons producing inhibitory gating of cutaneous sensory pathways during swimming and struggling.
Afferent Pathways↗
The neuronal targets for GABAergic reticulospinal inhibition that stops swimming in hatchling frog tadpoles.
In most animals locomotion can be started and stopped by specific sensory cues. We are using a simple vertebrate, the hatchling Xenopus tadpole, to study a neuronal pathway that turns off locomotion. In the tadpole, swimming stops when the head contacts solid objects or the water's surface meniscus. The primary sensory neurons are in the trigeminal ganglion and directly excite inhibitory reticulospinal neurons in the hindbrain. These project axons into the spinal cord and release GABA to inhibit spinal neurons and stop swimming. We ask whether there is specificity in the types of spinal neuron inhibited. We used single-neuron recording to determine which classes of spinal neurons receive inhibition when the head skin is pressed. Ventral motoneurons and premotor interneurons involved in generating the swimming rhythm receive reliable GABAergic inhibition. More dorsal inhibitory premotor interneurons are inhibited less reliably and some are excited. Dorsal sensory pathway interneurons that start swimming following a touch to the trunk skin do not appear to receive such inhibition. There is therefore specificity in the formation of descending inhibitory connections so that more ventral neurons producing swimming are most strongly inhibited.
Animals↗