Joel P. Gallagher

Metabotropic glutamate receptors are required for the induction of long-term potentiation

Recent observations have led to the suggestion that the metabotropic glutamate receptor may play a role in the induction or maintenance of long-term potentiation (LTP). However, experimental evidence supporting a role for this receptor in the induction of LTP is still inconclusive and controversial. Here we report that, in rat dorsolateral septal nucleus (DLSN) neurons, which have the highest density of metabotropic receptors and show functional responses, the induction of LTP is not blocked by the NMDA receptor antagonist 2-amino-5-phosphonovalerate, but is blocked by two putative metabotropic glutamate receptor antagonists, L-2-amino-3-phosphonopropionic acid and L-2-amino-4-phosphonobutyrate. Furthermore, superfusion of (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid, a selective metabotropic glutamate agonist, resulted in a long-lasting potentiation of synaptic transmission similar to that induced by tetanic stimuli. Our results demonstrated that activation of postsynaptic metabotropic receptors is both necessary and sufficient for the induction of LTP in the DLSN, and we suggest that such a mechanism may be important at other CNS synapses.

Biochemical and Electrophysiological Characteristics of Mammalian Gaba Receptors

The concept that GABA is a neurotransmitter in the mammalian CNS is supported by both electrophysiological and biochemical data. Whereas the electrophysiological studies are essential for demonstrating a specific functional response to GABA, the biochemical approach is useful for characterizing the molecular properties of this site. As a result of these studies the concept of the GABA receptor has progressed from a simple model of a single recognition site associated with a chloride channel to a more complex structure having a variety of interacting components. Thus, both electrophysiological and biochemical data support the existence of at least two pharmacologically distinct types of GABA receptors, based on the sensitivity to bicuculline. Also, anatomically, there appear to be two different types of receptors, those located postsynaptically on the soma or dendrites of a neighboring cell and those found presynaptically on GABAergic and other neurotransmitter terminals. From biochemical studies it appears that the GABA receptor may be composed of at least three distinct interacting components. One of these, the recognition site, may exist in two conformations, with one preferring agonists and the other having a higher affinity for antagonists. Ion channels may be considered a second component, with some of these regulating the passage of chloride ion, whereas others may be associated with calcium transport. The third major element of GABA receptors appears to be a benzodiazepine recognition site, although only a certain population of GABA receptors may be endowed with this property. In addition to these, the GABA receptor complex appears to contain substances that modulate the recognition site by influencing the availability of higher affinity binding proteins. It would appear therefore that changes affecting any one of these constituents can influence the characteristics of the others. While increasing the complexity of the system, this arrangement makes for a more sensitive and adaptable receptor mechanism. Thus the GABA receptor can be envisioned as a supramolecular complex of interacting sites, all of which contribute to the functional expression of receptor activation. Because of this complexity, GABA receptors can theoretically be modified in a variety of ways by drug treatment or disease. Accordingly, it may be possible to develop selective agonists and antagonists that may act at one of the basic components, as well as agents that may alter the receptor modulators. Conversely, a disorder of any of these entities may result in an alteration of GABA receptor function, which in turn could contribute to the symptoms of a variety of neuropsychiatric disorders.(ABSTRACT TRUNCATED AT 400 WORDS)

Corticotropin-Releasing Factor and Urocortin I Modulate Excitatory Glutamatergic Synaptic Transmission

Corticotropin-releasing factor (CRF)-related peptides serve as hormones and neuromodulators of the stress response and play a role in affective disorders. These peptides are known to alter complex behaviors and neuronal properties, but their receptor-mediated effects at CNS synapses are not well described. Here we show that excitatory glutamatergic transmission is modulated by two endogenous CRF-related peptide ligands, corticotropin-releasing factor [CRF rat/human (r/h)] and Urocortin I (Ucn I), within the central nucleus of the amygdala (CeA) and the lateral septum mediolateral nucleus (LSMLN). These limbic nuclei are reciprocally innervated, are involved in stress and affective disorders, and have high densities of the CRF receptors CRF1 and CRF2. Activation of these receptors exerts diametrically opposed actions on glutamatergic transmission in these nuclei. In the CeA, CRF(r/h) depressed excitatory glutamatergic transmission through a CRF1-mediated postsynaptic action, whereas Ucn I facilitated synaptic responses through presynaptic and postsynaptic CRF2-mediated mechanisms. Conversely, in the LSMLN, CRF caused a CRF1-mediated facilitation of glutamatergic transmission via postsynaptic mechanisms, whereas Ucn I depressed EPSCs by postsynaptic and presynaptic CRF2-mediated actions. Furthermore, antagonists of these receptors also affected glutamatergic neurotransmission, indicating that endogenous ligands tonically modulated synoptic activity at these synapses. These data show that CRF receptors in CeA and LSMLN synapses exert and maintain a significant synaptic tone and thereby regulate excitatory glutamatergic transmission. The results also suggest that CRF receptors may provide novel targets in affective disorders and stress.

Cocaine withdrawal enhances long‐term potentiation induced by corticotropin‐releasing factor at central amygdala glutamatergic synapses via CRF1, NMDA receptors and PKA

Cocaine addiction is an enduring, relapsing, behavioural disorder in which stressors reinstate cocaine‐seeking even after prolonged abstinence. Evidence suggests that the ‘anxiety‐like’ behaviour and stress associated with protracted withdrawal may be mediated by increased corticotropin‐releasing factor (CRF) in the central nucleus of the amygdala (CeA), a part of the limbic circuitry engaged in the coding and transmission of stimulus–reward associations. In the present study we describe a long‐lasting potentiation of glutamatergic transmission induced at lateral amygdala (LA)‐to‐CeA synapses by rat/human CRF. After 2 weeks of withdrawal from repeated intermittent exposure to cocaine, CRF‐induced long‐term potentiation (LTP) was greatly enhanced compared to the respective saline control group while, after short‐term withdrawal (24 h), there was no significant difference between the two treatment groups, indicating alterations in CRF systems during protracted withdrawal from chronic cocaine. After prolonged withdrawal, CRF‐induced LTP was dependent on activation of CRF2, CaV2.3 (R‐type) calcium channels and intracellular signalling through protein kinase C in both saline‐ and cocaine‐treated groups. The enhanced CRF‐induced LTP after 2 weeks of withdrawal was mediated through augmented CRF1 receptor function, associated with an increased signalling through protein kinase A, and required N‐methyl‐d‐aspartate (NMDA) receptors. Accordingly, single‐cell recordings revealed a significantly increased NMDA/AMPA ratio after prolonged withdrawal from the cocaine treatment. These results support a role for CRF1 receptor antagonists as plausible treatment options during withdrawal from chronic cocaine and suggest CaV2.3 blockers as potential candidates for pharmaceutical modulation of CRF systems.

Intracellular recordings from rat dorsolateral septal neurons, in vitro

We have made intracellular recordings from the dorsolateral septal nucleus (DLSN) in a rat brain slice. DLSN neurons fire short tetrodotoxin (TTX) sensitive action potentials followed by an after-hyperpolarization. The action potential is followed by either a Ca2+-dependent depolarization or a Ca2+-dependent after-hyperpolarization. Stimulation of medial septum results in antidromic invasion of DLSN neurons while stimulation of the fimbria/fornix results in orthodromic EPSPs or action potentials.

Comparison of antagonism by phaclofen of baclofen induced hyperpolarizations and synaptically mediated late hyperpolarizing potentials recorded intracellularly from rat dorsolateral septal neurons

Phaclofen has recently been described as an antagonist to baclofen at both peripheral and central receptors. We have applied phaclofen in known concentrations to an isolated rat brain slice preparation containing the septal nuclei. Our data demonstrate that phaclofen antagonizes responses to exogenously applied baclofen in a competitive manner. On the other hand, phaclofen is not as effective in antagonizing competitively the synaptically mediated late hyperpolarizing response (LHP) recorded from the same or similar dorsolateral septal nucleus (DLSN) neurons from which baclofen responses were recorded. Our data support the usefulness of phaclofen as a competitive antagonist of baclofen, and suggest that when larger stimulus intensities are applied, the LHP in the dorsolateral septum of the rat may be mediated by a transmitter in addition to gamma-aminobutyric acid (GABA).

The effects of temperature, pH and Cl-pump inhibitors on GABA responses recorded from cat dorsal root ganglia

GABA applied by iontophoresis produced GABA-induced currents (GCs) and GABA-induced depolarizations (GDs) which were recorded intracellularly from cat dorsal root ganglia (DRG). Lowering the temperature (37 to 27 degrees C) of the preparation depressed the amplitude of GCs while prolonging their rise-time and decay time. This depressant action was mainly due to a hyperpolarizing shift in the GABA equilibrium potential (EGABA). GABA responses could also be depressed by alkalinization of the superfusion solution or addition of putative chloride pump inhibitors, e.g. SITS, furosemide or bumetanide. However, the mechanism by which these latter procedures depressed GABA responses was not due to a shift in EGABA as occurred with lowered temperature. Instead we suggest that alkalinization or the putative chloride pump inhibitors affect the chloride channel or some other site associated with the GABA receptor complex and cause the depression we observed. GABA responses could be facilitated by lowering the pH of the superfusion solution or by injecting ammonium ion into a DRG. These results suggest that a temperature-sensitive, inwardly directed chloride pump that is resistant to SITS, furosemide or bumetanide, operates in cat DRG.

Chronic Cocaine Administration Switches Corticotropin-Releasing Factor2 Receptor-Mediated Depression to Facilitation of Glutamatergic Transmission in the Lateral Septum

Corticotropin-releasing factor (CRF) and urocortin (Ucn I) are endogenous members among a family of CRF-related peptides that activate two different and synaptically localized G-protein-coupled receptors, CRF1 and CRF2. These peptides and their receptors have been implicated in stress responses and stress with cocaine abuse. In this study, we observed significant alterations in excitatory transmission and CRF-related peptide regulation of excitatory transmission in the lateral septum mediolateral nucleus (LSMLN) after chronic cocaine administration. In brain slice recordings from the LSMLN of control (saline-treated) rats, glutamatergic synaptic transmission was facilitated by activation of CRF1 receptors with CRF but was depressed after activation of CRF2 receptors with Ucn I. After acute withdrawal from a chronic cocaine administration regimen, CRF1 activation remained facilitatory, but CRF2 activation facilitated rather than depressed LSMLN EPSCs. These alterations in CRF2 effects occurred through both presynaptic and postsynaptic mechanisms. In saline-treated rats, CRF1 and CRF2 coupled predominantly to protein kinase A signaling pathways, whereas after cocaine withdrawal, protein kinase C activity was more prominent and likely contributed to the CRF2-mediated presynaptic facilitation. Neither CRF nor Ucn I altered monosynaptic GABAA-mediated IPSCs before or after chronic cocaine administration, suggesting that loss of GABAA-mediated inhibition could not account for the facilitation. This switch in polarity of Ucn I-mediated neuromodulation, from a negative to positive regulation of excitatory glutamatergic transmission after chronic cocaine administration, could generate an imbalance in the brain reward circuitry associated with the LSMLN.

Histamine depolarizes rat medial vestibular nucleus neurons recorded intracellularly in vitro

The effects of histamine (HA) on the resting membrane potential and input resistance of rat medial vestibular nucleus (MVN) neurons were investigated using intracellular recording techniques from a submerged brain slice preparation. The exogenous application of HA predominantly produced a concentration-dependent membrane depolarization and induction of action potential firing. The depolarization exhibited a rapid onset, a slow recovery, and usually occurred in the absence of any apparent change in conductance. These effects of HA could be mimicked by the H2-agonist impromidine and were reversibly blocked by the H2-antagonist cimetidine. Tetrodotoxin (TTX) or low calcium/high magnesium-containing media failed to block completely the HA-induced depolarization supporting a direct postsynaptic receptor mediated action of HA. The diminished HA-induced depolarization observed following pretreatment with TTX cannot exclude an additional presynaptic action by HA. The present findings reveal that HA exerts a novel direct excitation of rat MVN neurons through an H2-receptor.

Modulation of Excitatory Transmission at the Rat Medial Vestibular Nucleus Synapsea

Information transfer from the peripheral vestibular apparatus to the central vestibular nuclear complex (VNC) and the intrinsic processing of this information within the VNC must first be understood in order to define normal vestibular function and the etiology of a variety of clinical vestibular disorders. It is only when the specific types of receptors responsible for the cellular chemical transduction at synapses within this complex are identified and characterized that an optimal rational therapeutic approach towards treating vestibular disorders will be realized. As a first step towards achieving this goal we have developed an in vitro brain slice preparation from rats’.’ that contains components of the central VNC, specifically, the primary vestibular nerve (N. VIII) afferent inputs and second-order neurons in the medial and lateral vestibular nuclei (FIGURE 1A). We have coupled electrophysiological, primarily intracellular, recording techniques with this in vitro brain slice preparation to examine the cellular pharmacology at the primary afferent to second-order synapse in the medial vestibular nucleus (MVN). The development of the in vitro brain slice preparation has allowed us to minimize many of the difficulties associated commonly with attempts to record intracellularly from the VNC in the intact animal. On the other hand, by reducing the preparation to the central components of the vestibular system, we have unfortunately removed all of the receptor-driven inputs that the brain normally receives from the peripheral vestibular apparatus as well as the extrinsic circuit inputs arising from other descending and ascending influences that are active in an intact animal. Nonetheless, we are still able to evoke “typical” excitatory synaptic transmission with our isolated in vitro preparation by stimulating the primary vestibular afferent fibers in the N.VIII rootlet and along its pathway to the MVN (FIGURE 1B). Furthermore, it is apparent that other nerve terminals (e.g., GABAergic) remain functionally intact in the slice and are fully capable of releasing their transmitters, since application of selective antagonists for various substances modifies spontaneous electrical and synaptic activity. A significant finding associated with the development of the in vitro preparation was the observation that MVN neurons exhibited spontaneous action potentials,’“ i.e., they were not quiescent, despite the fact that they were no longer under the