Local Inhibitory Inputs Research Articles

Differential wiring of local excitatory and inhibitory synaptic inputs to islet cells in rat spinal lamina II demonstrated by laser scanning photostimulation

The substantia gelatinosa (lamina II) of the spinal dorsal horn contains inhibitory and excitatory interneurons that are thought to play a critical role in the modulation of nociception. However, the organization of the intrinsic circuitry within lamina II remains poorly understood. We used glutamate uncaging by laser scanning photostimulation to map the location of neurons that give rise to local synaptic inputs to islet cells, a major class of inhibitory interneuron in lamina II. We also mapped the distribution of sites on the islet cells that exhibited direct (non-synaptic) responses to uncaging of excitatory and inhibitory transmitters. Local synaptic inputs to islet cells arose almost entirely from within lamina II, and these local inputs included both excitatory and inhibitory components. Furthermore, there was a striking segregation in the location of sites that evoked excitatory versus inhibitory synaptic inputs, such that inhibitory presynaptic neurons were distributed more proximal to the islet cell soma. This was paralleled in part by a differential distribution of transmitter receptor sites on the islet cell, in that inhibitory sites were confined to the peri-somatic region while excitatory sites were more widespread. This differential organization of excitatory and inhibitory inputs suggests a principle for the wiring of local circuitry within the substantia gelatinosa.

Local Generation of Theta-Frequency EEG Activity in the Parasubiculum

The parasubiculum is a major component of the hippocampal formation that receives inputs from the CA1 region, anterior thalamus, and medial septum and that projects primarily to layer II of the entorhinal cortex. Hippocampal theta-frequency (4-12 Hz) electroencephalographic (EEG) activity has been correlated with sensorimotor integration, spatial navigation, and memory functions. The present study was aimed at determining if theta is also generated locally within the parasubiculum versus volume conducted from adjacent structures. In urethan-anesthetized rats, the phase-reversal of theta activity between superficial and deep layers of the parasubiculum was demonstrated using differential recordings from movable bipolar electrodes that eliminate the influence of volume-conducted activity. Parasubicular theta was abolished by atropine, and was in phase with theta in stratum radiatum/lacunosum-moleculare of the CA1 region. Whole cell current-clamp recordings in brain slices were then used to determine if parasubicular theta may be generated in part by membrane potential oscillations in layer II neurons. Membrane potential oscillations occurred in most layer II neurons, including four putative interneurons, when cells were held at near-threshold voltages using current injection. The frequency of oscillations increased from 3.2 to 6.1 Hz when bath temperature was raised from 22 to 32 degrees C, and oscillations persisted in the presence of blockers of fast ionotropic glutamatergic and GABAergic synaptic transmission. Oscillations are therefore likely generated by intrinsic, voltage-dependent ionic conductances. These results indicate that theta field activity is generated locally within the parasubiculum and that intrinsic membrane potential oscillations, synchronized by local inhibitory inputs, may contribute to the generation of this activity.

GABAB Receptor Isoforms Caught in Action at the Scene

The metabotropic GABAB receptors mediate slow synaptic inhibition and consist of heterodimers of GABAB1 and GABAB2 subunits. The only known molecular diversity of the GABAB receptors arises from the two GABAB1 isoforms, but its functional significance has been unclear. Two studies in this issue of Neuron now demonstrate that GABAB1a and GABAB1b show strategically distinct subcellular localization and physiological action.

The GABAB1b Isoform Mediates Long-Lasting Inhibition of Dendritic Ca2+ Spikes in Layer 5 Somatosensory Pyramidal Neurons

The apical tuft of layer 5 pyramidal neurons is innervated by a large number of inhibitory inputs with unknown functions. Here, we studied the functional consequences and underlying molecular mechanisms of apical inhibition on dendritic spike activity. Extracellular stimulation of layer 1, during blockade of glutamatergic transmission, inhibited the dendritic Ca2+ spike for up to 400 ms. Activation of metabotropic GABAB receptors was responsible for a gradual and long-lasting inhibitory effect, whereas GABAA receptors mediated a short-lasting (approximately 150 ms) inhibition. Our results suggest that the mechanism underlying the GABAB inhibition of Ca2+ spikes involves direct blockade of dendritic Ca2+ channels. By using knockout mice for the two predominant GABAB1 isoforms, GABAB1a and GABAB1b, we showed that postsynaptic inhibition of Ca2+ spikes is mediated by GABAB1b, whereas presynaptic inhibition of GABA release is mediated by GABAB1a. We conclude that the molecular subtypes of GABAB receptors play strategically different physiological roles in neocortical neurons.

Direction-Selective Dendritic Action Potentials in Rabbit Retina

Dendritic spikes that propagate toward the soma are well documented, but their physiological role remains uncertain. Our in vitro patch-clamp recordings and two-photon calcium imaging show that direction-selective retinal ganglion cells (DSGCs) utilize orthograde dendritic spikes during physiological activity. DSGCs signal the direction of image motion. Excitatory subthreshold postsynaptic potentials are observed in DSGCs for motion in all directions and provide a weakly tuned directional signal. However, spikes are generated over only a narrow range of motion angles, indicating that spike generation greatly enhances directional tuning. Our results indicate that spikes are initiated at multiple sites within the dendritic arbors of DSGCs and that each dendritic spike initiates a somatic spike. We propose that dendritic spike failure, produced by local inhibitory inputs, might be a critical factor that enhances directional tuning of somatic spikes.

Synaptic integration in motoneurons with hyper-excitable dendrites

Motoneurons have extensive dendritic trees that receive the numerous inputs required to produce movement. These dendrites are highly active, containing voltage-sensitive channels that generate persistent inward currents (PICs) that can enhance synaptic input 5-fold or more. However, this enhancement is proportional to the level of activity of monoaminergic inputs from the brainstem that release serotonin and noradrenalin. The higher this activity, the larger the dendritic PIC and the higher the firing rate evoked by a given amount of excitatory synaptic input. This brainstem control of motoneuron input-output gain translates directly into control of system gain of a motor pool and its muscle. Because large dendritic PICs are probably necessary for motoneurons to have sufficient gain to generate large forces, it is possible that descending monoaminergic inputs scale in proportion to voluntary force. Inhibition from sensory inputs has a strong suppressive effect on dendritic PICs: the stronger the inhibition, the smaller the PIC. Thus, local inhibitory inputs within the cord may oppose the descending monoaminergic control of PICs. Most motor behaviors evoke a mixture of excitation and inhibition (e.g., the reciprocal inhibition between antagonists). Therefore, normal joint movements may involve constant adjustment of PIC amplitude.

Activation of presynaptic glycine receptors facilitates glycine release from presynaptic terminals synapsing onto rat spinal sacral dorsal commissural nucleus neurons.

Glycine is a major inhibitory neurotransmitter in the spinal cord and brainstem. Here we report the novel finding that presynaptic glycine autoreceptors modulate release from terminals synapsing onto rat spinal sacral dorsal commissural nucleus (SDCN) neurons. In mechanically dissociated SDCN neurons, in which functional presynaptic nerve terminals remain adherent to the isolated neurons, exogenously applied glycine (3 microM) increased the frequency of glycinergic spontaneous inhibitory postsynaptic currents (sIPSCs) without affecting their amplitudes or decay times. This suggests that glycine acts presynaptically to increase glycine release probability. Picrotoxin, at a concentration that had little direct effect on sIPSC frequency and amplitude (30 microM), significantly attenuated glycine-induced presynaptic sIPSC facilitation. The glycine-induced sIPSC frequency facilitation was completely abolished either in a Ca(2+)-free external solution or in the presence of 100 microM Cd2+, suggesting the involvement of extracellular Ca2+ influx into the nerve terminals. The glycine action was also completely occluded in the presence of 300 nM tetrodotoxin. In recordings from SDCN neurons in spinal cord slices, glycine (10 microM) increased evoked IPSC (eIPSC) amplitude and decreased the extent of paired-pulse facilitation. In response to brief high frequency stimulus trains the eIPSCs displayed a profound frequency-dependent facilitation that was greatly reduced by picrotoxin (30 microM). These results indicate that glycine acts at presynaptic autoreceptors, causing depolarization of the glycinergic nerve terminals, the subsequent activation of voltage-dependent Na+ and Ca2+ channels, and facilitation of glycine release. Furthermore, this presynaptic facilitation was observed under more physiological conditions, suggesting that these glycinergic autoreceptors may contribute to the integration of local inhibitory inputs to SDCN neurons.

The role of the thalamus in the flow of information to the cortex.

The lateral geniculate nucleus is the best understood thalamic relay and serves as a model for all thalamic relays. Only 5-10% of the input to geniculate relay cells derives from the retina, which is the driving input. The rest is modulatory and derives from local inhibitory inputs, descending inputs from layer 6 of the visual cortex, and ascending inputs from the brainstem. These modulatory inputs control many features of retinogeniculate transmission. One such feature is the response mode, burst or tonic, of relay cells, which relates to the attentional demands at the moment. This response mode depends on membrane potential, which is controlled effectively by the modulator inputs. The lateral geniculate nucleus is a first-order relay, because it relays subcortical (i.e. retinal) information to the cortex for the first time. By contrast, the other main thalamic relay of visual information, the pulvinar region, is largely a higher-order relay, since much of it relays information from layer 5 of one cortical area to another. All thalamic relays receive a layer-6 modulatory input from cortex, but higher-order relays in addition receive a layer-5 driver input. Corticocortical processing may involve these corticothalamocortical 're-entry' routes to a far greater extent than previously appreciated. If so, the thalamus sits at an indispensable position for the modulation of messages involved in corticocortical processing.

Hypocretins (Orexins) Regulate Serotonin Neurons in the Dorsal Raphe Nucleus by Excitatory Direct and Inhibitory Indirect Actions

The hypocretins (hcrt1 and hcrt2) are expressed by a discrete population of hypothalamic neurons projecting to many regions of the CNS, including the dorsal raphe nucleus (DRN), where serotonin (5-HT) neurons are concentrated. In this study, we investigated responses to hcrts in 216 physiologically identified 5-HT and non-5-HT neurons of the DRN using intracellular and whole-cell recording in rat brain slices. Hcrt1 and hcrt2 induced similar amplitude and dose-dependent inward currents in most 5-HT neurons tested (EC50, approximately 250 nm). This inward current was not blocked by the fast Na+ channel blocker TTX or in a Ca2+-free solution, indicating a direct postsynaptic action. The hcrt-induced inward current reversed near -18 mV and was primarily dependent on external Na+ but not on external or internal Ca2+, features typical of Na+/K+ nonselective cation channels. At higher concentrations, hcrts also increased spontaneous postsynaptic currents in 5-HT neurons (EC50, approximately 450-600 nm), which were TTX-sensitive and mostly blocked by the GABA(A) antagonist bicuculline, indicating increased impulse flow in local GABA interneurons. Accordingly, hcrts were found to increase the basal firing of presumptive GABA interneurons. Immunolabeling showed that hcrt fibers projected to both 5-HT and GABA neurons in the DRN. We conclude that hcrts act directly to excite 5-HT neurons primarily via a TTX-insensitive, Na+/K+ nonselective cation current, and indirectly to activate local inhibitory GABA inputs to 5-HT cells. The greater potency of hcrts in direct excitation compared with indirect inhibition suggests a negative feedback function for the latter at higher levels of hcrt activity.

Chapter 4 Thalamic relay functions

The lateral geniculate nucleus is the best understood thalamic relay. Only 5-10% of the inputs to geniculate relay cells derive from retina, which is the driving input. The rest, being modulatory, derive from local inhibitory inputs, descending inputs from visual cortex, and ascending inputs from brainstem. The nonretinal, modulatory inputs, which form the vast majority, dynamically control the nature of the geniculate relay. Among other actions, these modulatory inputs regulate membrane properties of relay cells and thereby control their mode of response to retinal inputs, and this dramatically affects the nature of information relayed to cortex. Our studies of the lateral geniculate nucleus of the cat lead to the speculation that this dynamic control depends on the animal's behavioral state and represents the neuronal substrate for many forms of visual attention. The lateral geniculate nucleus is a first-order relay, because it relays subcortical (i.e. retinal) information to cortex for the first time. In contrast, the other main thalamic relay of visual information, the pulvinar (and lateral posterior nucleus in carnivores), is largely a higher-order relay, since much of it seems to relay information from one cortical area to another. Much more corticocortical processing may involve these 're-entry' routes than has been hitherto appreciated. If so, the thalamus sits at an indispensable position for corticocortical processing.

Organization of Intracortical Circuits in Relation to Direction Preference Maps in Ferret Visual Cortex

Neurons in the primary visual cortex are selective for the direction of movement of a visual stimulus. Like other stimulus features, direction preference is mapped on the cortical surface in a systematic manner. Intracortical synaptic circuits, in particular inhibitory connections, have been implicated in the emergence of direction selectivity. Whether intracortical inhibition specifically suppresses responses to the nonpreferred direction or has a nonspecific "thresholding" effect is still controversial. To address these questions we investigated the relationship between patterns of intracortical synaptic connections and direction domains in ferret primary visual cortex (area 17) using a combined in vivo-in vitro approach. Excitatory synaptic inputs were iso-direction-tuned. The majority of local inhibitory inputs were also iso-direction-tuned. However, approximately 40% of inhibitory connections originated in regions preferring the opposite direction. These findings indicate that specific inhibitory interactions between cortical regions of opposite direction preference may contribute to the emergence and sharpening of direction selectivity.

Physiological Mapping of Local Inhibitory Inputs to the Hypothalamic Paraventricular Nucleus

Local inhibitory synaptic inputs to neurons of the rat hypothalamic paraventricular nucleus (PVN) were studied by using glutamate microstimulation and conventional intracellular and whole-cell patch-clamp recording in coronal, horizontal, and parasagittal slices of rat hypothalamus. PVN cells were classified as magnocellular or parvocellular neurons on the basis of electrophysiological and post hoc immunohistochemical analyses; GABA-producing neurons were localized with in situ hybridization. Glutamate microstimulation of different sites around the PVN evoked volleys of postsynaptic potentials in 43% of the PVN neurons tested. Some responses to stimulation at each site were blocked by bicuculline, suggesting that they were mediated by the activation of presynaptic GABA neurons. In the coronal plane, presynaptic inhibitory sites were located lateral to the PVN and ventral to the fornix, corresponding to the lateral hypothalamic area and the posterior bed nucleus of the stria terminalis (BNST). In the horizontal plane, presynaptic inhibitory sites were found rostral, lateral, and caudal to the nucleus, corresponding to parts of the anterior hypothalamic area, the posterior BNST, the medial preoptic area, and the dorsomedial hypothalamus. In the parasagittal plane, presynaptic inhibitory neurons were revealed at sites rostral and caudal to the nucleus, corresponding to the medial preoptic area and the dorsomedial hypothalamus, and in a site dorsal to the optic chiasm that included the suprachiasmatic nucleus. These presynaptic sites each contained GABA-producing neurons based on in situ hybridization with a glutamic acid decarboxylase riboprobe and together formed a three-dimensional ring around the PVN. Unexpectedly, both magnocellular and parvocellular neurons received inhibitory synaptic inputs from common sites.

Brain-stem modulation of the response properties of cells in the cat's perigeniculate nucleus.

Transmission through the lateral geniculate nucleus is facilitated following activation of the cholinergic input from the brain stem, which is thought to reflect activity patterns seen during arousal. One of the underlying mechanisms is the suppression of inhibitory circuits local to the lateral geniculate nucleus. However, evidence exists that some visually driven inhibitory inputs to geniculate nucleus. preserved or even enhanced under conditions of arousal, and during electrical activation of the parabrachial region of the brain stem. We have therefore reexamined the effect of brain-stem activation on the visual responses of one group of local inhibitory inputs to geniculate relay cells, those emanating from the adjacent perigeniculate nucleus. We recorded single perigeniculate cells in anesthetized, paralyzed cats. Axons innervating the lateral geniculate and perigeniculate nuclei from the parabrachial region of the brain stem were electrically activated, and the effect of this activation was assessed on both spontaneous and visually evoked responses. Visual stimulation consisted of sinusoidally modulated sine-wave gratings of varying spatial and temporal frequency. For the great majority of perigeniculate cells (32 of 40), brain-stem activation inhibited spontaneous activity, while one cell was excited, three showed a mixed effect and four were unaffected. Nevertheless, the responses of most cells (30 of 40) were facilitated when brain-stem activation was paired with certain spatio-temporal patterns of visual stimulation. Spatial tuning curves were constructed for 17 cells and temporal tuning curves for 14, before and during parabrachial activation. The responses of any one cell could be facilitated, unchanged, or suppressed, depending on the visual stimulus used. In some cases, this substantially modified the cell's spatial and temporal tuning properties. We conclude that activation of the brain stem disinhibits geniculate relay cells in the absence of visual stimulation, but it has the potential to enhance either the magnitude or specificity of visually driven inhibition arising from the perigeniculate nucleus.

Local inhibitory synaptic inputs to neurones of the paraventricular nucleus in slices of rat hypothalamus.

1. Intracellular recordings were obtained from neurones in the region of the paraventricular nucleus in slices of rat hypothalamus. Glutamate microdrops were applied to the surface of the slices at sites dorsal, lateral and ventral to the paraventricular nucleus to selectively activate local presynaptic neurones. The gamma-aminobutyric acidA (GABAA)-receptor antagonists picrotoxin or bicuculline were bath-applied to block synaptic inhibition. 2. Glutamate microapplication caused a tonic depolarization and often repetitive action potentials in twenty of forty-seven recorded cells. This was probably caused by the direct exposure of the dendrites of the recorded cells to the glutamate microdrops. 3. Glutamate microstimulation elicited inhibitory synaptic responses in nine of forty-seven neurones tested. Glutamate microdrops caused discrete, hyperpolarizing postsynaptic potentials (PSPs) in four cells recorded with microelectrodes containing potassium acetate and evoked depolarizing PSPs in four cells recorded with KCl-filled microelectrodes. Glutamate microapplication inhibited spontaneous spike firing in another cell recorded with a potassium acetate microelectrode. 4. Bath application of GABAA-receptor antagonists completely blocked the hyperpolarizing PSPs elicited by glutamate microstimulation in three of three cells recorded with potassium acetate electrodes and the depolarizing PSPs in two of two cells recorded with KCl electrodes, indicating they were inhibitory PSPs caused by the release of GABA. Suppression of GABAA-mediated synaptic inhibition did not reveal any glutamate-evoked excitatory PSPs. 5. Recorded cells were identified as magnocellular, parvocellular or non-paraventricular bursting neurones on the basis of their electrophysiological properties. Direct depolarization and local inhibitory synaptic responses were observed in all three cell types. 6. Several conclusions can be drawn from these data: (1) functional glutamate receptors are distributed throughout neuronal populations in the paraventricular region of the hypothalamus, confirming and extending previous observations; (2) local synaptic inputs to neurones in the paraventricular nucleus are primarily inhibitory, supplied by perinuclear GABAergic neurones; (3) both magnocellular and parvocellular subpopulations receive local inhibitory synaptic inputs. The possibility that these local GABAergic circuits mediate inhibitory inputs to paraventricular neurones from limbic structures is discussed.