Kimmo Jensen

Hippocampal GABAergic dysfunction in a rat chronic mild stress model of depression

In major depression, one line of research indicates that a dysfunctional GABAergic inhibitory system is linked to the appearance of depressive symptoms. However, as the mechanistic details of such GABAergic deficit are largely unknown, we undertook a functional investigation of the GABAergic system in the rat chronic mild stress model of depression. Adult rats were exposed to an eight‐week long stress protocol leading to anhedonic‐like behavior. In hippocampal brain slices, phasic, and tonic GABAA receptor‐mediated currents in dentate gyrus granule cells were examined using patch‐clamp recordings. In granule cells, the frequency of spontaneous inhibitory postsynaptic currents (sIPSCs) was reduced to 41% in anhedonic‐like rats, which was associated with a reduced probability of evoked GABA release. Using immunohistochemical analysis, there was no change in the number of parvalbumin‐positive interneurons in the dentate gyrus. Notably, we observed a 60% increase in THIP‐activated tonic GABAA mediated current in anhedonic‐like rats, suggesting an upregulation of extrasynaptic GABAA receptors. Finally, five weeks treatment with the antidepressant escitalopram partially reversed the sIPSCs frequency. In summary, we have revealed a hippocampal dysfunction in the GABAergic system in the chronic mild stress model of depression in rats, caused by a reduction in action potential‐dependent GABA release. Since the function of the GABAergic system was improved by antidepressant treatment, in parallel with behavioral read outs, it suggests a role of the GABAergic system in the pathophysiology of depression.

Mature BDNF, But Not proBDNF, Reduces Excitability of Fast-Spiking Interneurons in Mouse Dentate Gyrus

Mature BDNF and its precursor proBDNF may both be secreted to exert opposite effects on synaptic plasticity in the hippocampus. However, it is unknown how proBDNF and mature BDNF affect the excitability of GABAergic interneurons and thereby regulate GABAergic inhibition. We made recordings of GABAergic spontaneous IPSCs (sIPSCs) in mouse dentate gyrus granule cells and found that chronic or acute BDNF reductions led to large increases in the sIPSC frequencies, which were TTX (tetrodotoxin) sensitive and therefore action-potential driven. Conversely, addition of mature BDNF, but not proBDNF, within minutes led to a decrease in the sIPSC frequency to 44%. Direct recordings from fast-spiking GABAergic interneurons revealed that mature BDNF reduced their excitability and depressed their action potential firing, whereas proBDNF had no effect. Using the TrkB inhibitor K-252a, or mice deficient for the common neurotrophin receptor p75NTR, the regulation of GABAergic activity was shown specifically to be mediated by BDNF binding to the neurotrophin receptor TrkB. In agreement, immunohistochemistry demonstrated that TrkB, but not p75NTR, was expressed in parvalbumin-positive interneurons. Our results suggest that mature BDNF decreases the excitability of GABAergic interneurons via activation of TrkB, while proBDNF does not impact on GABAergic activity. Thus, by affecting the firing of GABAergic interneurons, mature BDNF may play an important role in regulating network oscillations in the hippocampus.

Post‐tetanic potentiation of GABAergic IPSCs in cultured rat hippocampal neurones

1 Dual whole‐cell patch‐clamp recording was used to investigate post‐tetanic potentiation (PTP) of GABAergic IPSCs evoked between pairs of cultured rat hippocampal neurones. Tetanization of the presynaptic neurone at frequencies (f) ranging from 5 to 100 Hz resulted in PTP of the IPSCs. Maximum PTP had a magnitude of 51.6 % just after the stimulus train, and lasted up to 1 min. PTP was shown to be dependent on the number of stimuli in the train, but independent of f at frequencies ≥ 5 Hz. 2 Blocking postsynaptic GABAA receptors with bicuculline during the tetanus did not affect the expression of PTP, showing that it is a presynaptic phenomenon. PTP was strongly affected by changing [Ca2+]oduring the tetanus: PTP was reduced by lowering [Ca2+]o, and increased by high [Ca2+]o. 3 PTP was still present after presynaptic injection of BAPTA or EGTA, or following perfusion of the membrane‐permeable ester EGTA‐tetraacetoxymethyl ester (EGTA AM, 50 μM). On the other hand, EGTA AM blocked spontaneous, asynchronous IPSCs (asIPSCs), which were often associated with tetanic stimulation. 4 Tetanic stimulation in the presence of 4‐aminopyridine (4‐AP), which promotes presynaptic Ca2+ influx, evoked sustained PTP of IPSCs in half of the neurones tested. 5 The results indicate that PTP at inhibitory GABAergic synapses is related to the magnitude of presynaptic Ca2+ influx during the tetanic stimulation, leading to an enhanced probability of vesicle release in the post‐tetanic period. The increase in [Ca2+]i occurs despite the presence of high‐affinity exogenous and endogenous intracellular Ca2+ buffers. That PTP of IPSCs depends on the number, and not the frequency, of spikes in the GABAergic neurone is in accordance with a slow clearing of intracellular Ca2+ from the presynaptic terminals.

Cell Type–Specific GABAA Receptor–Mediated Tonic Inhibition in Mouse Neocortex

Activity of extrasynaptic GABA A receptors mediating tonic inhibition is thought to play an important role for the excitability of the mammalian cerebral cortex. However, little is known about the cell type-specific expression of tonic inhibition in particular types of cortical interneurons. Here, we used transgenic mice expressing green fluorescent protein (GFP) in somatostatin-positive (SOM) interneurons and investigated tonic inhibition in SOM interneurons versus pyramidal cells in neocortical layers 2/3. In brain slices, pyramidal cells showed a tonic current of 66 +/- 19 pA in response to the delta-subunit selective GABA A agonist THIP (1 microM). On the other hand, tonic inhibition was absent in SOM interneurons (8 +/- 1 pA) in response to THIP. As opposed to pyramidal cells, SOM interneurons were also insensitive to the delta-subunit preferring neurosteroid allotetrahydrodeoxycorticosterone (THDOC) (100 nM) and to elevated endogenous GABA levels in the slice. Finally, SOM interneurons received only 45% of the phasic charge transfer during GABA A receptor-mediated synaptic activity compared with pyramidal cells. Altogether, our study indicates that SOM interneurons receive relatively weak inhibitory input and cannot be brought under the influence of tonic inhibition.

Reduced GABAergic Inhibition Explains Cortical Hyperexcitability in the Wobbler Mouse Model of ALS

Amyotrophic lateral sclerosis (ALS) is a progressive degenerative disease of the central nervous system. Symptomatic and presymptomatic ALS patients demonstrate cortical hyperexcitability, which raises the possibility that alterations in inhibitory gamma-aminobutyric acid (GABA)ergic system could underlie this dysfunction. Here, we studied the GABAergic system in cortex using patch-clamp recordings in the wobbler mouse, a model of ALS. In layer 5 pyramidal neurons of motor cortex, the frequency of GABA(A) receptor-mediated spontaneous inhibitory postsynaptic currents was reduced by 72% in wobbler mice. Also, miniature inhibitory postsynaptic currents recorded under blockade of action potentials were decreased by 64%. Tonic inhibition mediated by extrasynaptic GABA(A) receptors was reduced by 87%. In agreement, we found a decreased density of parvalbumin- and somatostatin-positive inhibitory interneurons and reduced vesicular GABA transporter immunoreactivity in the neuropil. Finally, we observed an increased input resistance and excitability of wobbler excitatory neurons, which could be explained by lack of GABA(A) receptor-mediated influences. In conclusion, we demonstrate decreases in GABAergic inhibition, which might explain the cortical hyperexcitability in wobbler mice.

Pharmacological characterization of a novel positive modulator at α4β3δ-containing extrasynaptic GABAA receptors

The in vitro and in vivo pharmacological effects of [2-amino-4-(2,4,6-trimethylbenzylamino)-phenyl]-carbamic acid ethyl ester (AA29504), which is a close analogue of retigabine, have been investigated. AA29504 induced a rightward shift of the activation threshold at cloned KCNQ2, 2/3 and 4 channels expressed in Xenopus oocytes, with a potency 3-4fold lower than retigabine. AA29504 (1 muM) had no agonist activity when tested at alpha(1)beta(3)gamma(2s) or alpha(4)beta(3)delta GABA(A) receptors expressed in Xenopus oocytes, but left-shifted the EC(50) for GABA and gaboxadol (THIP) at both receptors. The maximum GABA response at alpha(1)beta(3)gamma(2s) receptors was unchanged by AA29504 (1 muM), but increased 3-fold at alpha(4)beta(3)delta receptors. In slices prepared from the prefrontal cortex of adult rats AA29504 had no effect alone on the average IPSC or the tonic current in layer II/III pyramidal neurons, but potentiated the effect of gaboxadol on both phasic and tonic currents. Thus, the effects of gaboxadol could be positively modulated by AA29504. Systemic administration of AA29504 at doses relevant for modulating GABA transmission produced anxiolytic effects and reduced motor coordination consistent with activity at GABA(A) receptors. We conclude that AA29504 exerts a major action via alpha(4)beta(3)delta-containing GABA(A) receptors, which will be important for interpreting its effect in vivo.

Selective sparing of hippocampal CA3 cells following in vitro ischemia is due to selective inhibition by acidosis

A brief global ischemic insult to the brain leads to a selective degeneration of the pyramidal neurons in the hippocampal CA1 region while the neurons in the neighbouring CA3 region are spared. The reason for this difference is not known. The selective vulnerability of CA1 neurons to ischemia can be reproduced in vitro in murine organotypic slice cultures, if the ion concentrations in the medium during the anoxic/aglycemic insult are similar to that in the brain extracellular fluid during ischemia in vivo. As acidosis develops during ischemia, we studied the importance of extracellular pH for selective vulnerability. We found that cell death in the CA1 and CA3 regions was equally prevented by removal of calcium from the medium or following blockade of the N‐methyl‐d‐aspartate (NMDA) receptor by d‐2 amino‐5‐phosphonopentanoic‐acid (D‐APV). On the other hand, damage to the CA3 neurons markedly decreased with decreasing pH following in vitro ischemia, while the degeneration of CA1 neurons was less pH dependent. Patch‐clamp recordings from pyramidal neurons in the CA1 and CA3 regions, respectively, revealed a pronounced inhibition of NMDA‐receptor mediated excitatory postsynaptic currents (EPSCs) at pH 6.5 that was equally pronounced in the two regions. However, when changing pH from 6.5 to 7.4 the recovery of the EPSCs was significantly slower in the CA3 region. We conclude that acidosis selectively protects CA3 pyramidal neurons during in vitro ischemia, and differentially affects the kinetics of NMDA receptor activation, which may explain the difference in vulnerability between CA1 and CA3 pyramidal neurons to an ischemic insult.

SorCS2 is required for BDNF -dependent plasticity in the hippocampus

SorCS2 is a member of the Vps10p-domain receptor gene family receptors with critical roles in the control of neuronal viability and function. Several genetic studies have suggested SORCS2 to confer risk of bipolar disorder, schizophrenia and attention deficit-hyperactivity disorder. Here we report that hippocampal N-methyl-d-aspartate receptor-dependent synaptic plasticity is eliminated in SorCS2-deficient mice. This defect was traced to the ability of SorCS2 to form complexes with the neurotrophin receptor p75NTR, required for pro-brain-derived neurotrophic factor (BDNF) to induce long-term depression, and with the BDNF receptor tyrosine kinase TrkB to elicit long-term potentiation. Although the interaction with p75NTR was static, SorCS2 bound to TrkB in an activity-dependent manner to facilitate its translocation to postsynaptic densities for synaptic tagging and maintenance of synaptic potentiation. Neurons lacking SorCS2 failed to respond to BDNF by TrkB autophosphorylation, and activation of downstream signaling cascades, impacting neurite outgrowth and spine formation. Accordingly, Sorcs2–/– mice displayed impaired formation of long-term memory, increased risk taking and stimulus seeking behavior, enhanced susceptibility to stress and impaired prepulse inhibition. Our results identify SorCS2 as an indispensable coreceptor for p75NTR and TrkB in hippocampal neurons and suggest SORCS2 as the link between proBDNF/BDNF signaling and mental disorders.

SSADH deficiency leads to elevated extracellular GABA levels and increased GABAergic neurotransmission in the mouse cerebral cortex

SummarySuccinic semialdehyde dehydrogenase (SSADH) deficiency is an inherited disorder in which patients display neurodevelopmental retardation, ataxia, and epileptic seizures. The recently engineered SSADH knock-out (KO) mouse models the severe form of the human disorder. The SSADH enzyme participates in the breakdown of the inhibitory neurotransmitter GABA, and studies have shown increases in brain GABA and downregulation of GABAA receptor β2 subunits in the cerebral cortex of these mice. Here, we used brain slice electrophysiology to investigate the alterations in GABA neurotransmission in SSADH KO mouse cortex. In layer 2/3 pyramidal cells, spontaneous inhibitory postsynaptic currents (IPSCs), reflecting activity of GABAergic synaptic contacts, were normal in SSADH KO mice. Also, IPSCs evoked by electrical single-axon stimulation in KO mice were normal. In contrast, tonic inhibition mediated by presumed extrasynaptic GABAA receptors was strongly increased, indicating significantly raised extracellular GABA levels. The excessive cortical GABAergic neurotransmission may participate in the seizure activity in SSADH deficiency.

BDNF Depresses Excitability of Parvalbumin-Positive Interneurons through an M-Like Current in Rat Dentate Gyrus

In addition to their classical roles in neuronal growth, survival and differentiation, neurotrophins are also rapid regulators of excitability, synaptic transmission and activity-dependent synaptic plasticity. We have recently shown that mature BDNF (Brain Derived Neurotrophic Factor), but not proBDNF, modulates the excitability of interneurons in dentate gyrus within minutes. Here, we used brain slice patch-clamp recordings to study the mechanisms through which BDNF modulates the firing of interneurons in rat dentate gyrus by binding to TrkB receptors. Bath application of BDNF (15 ng/ml) under current-clamp decreased the firing frequency (by 80%) and input resistance, blocking the delayed firing observed at near-threshold voltage ranges, with no changes in resting membrane potential or action potential waveform. Using TEA (tetraethylammonium), or XE991(a Kv7/KCNQ channel antagonist), the effect of BDNF was abolished, whereas application of retigabine (a Kv7/KCNQ channel opener) mimicked the effect of BDNF, suggesting that the M-current could be implicated in the modulation of the firing. In voltage-clamp experiments, BDNF increased the M-like current amplitude with no change in holding current. This effect was again blocked by XE991 and mimicked by retigabine, the latter accompanied with a change in holding current. In agreement with the electrophysiology, parvalbumin-positive interneurons co-expressed TrkB receptors and Kv7.2/KCNQ2 channels. In conclusion, BDNF depresses the excitability of interneurons by activating an M-like current and possibly blocking Kv1 channels, thereby controlling interneuron resting membrane potential and excitability.