Different input and output properties characterize parvalbumin‐positive basket and Axo‐axonic cells in the hippocampal CA3 subfield

In the hippocampus, parvalbumin‐expressing basket (BC) and axo‐axonic cells (AAC) show different discharge patterns during distinct network states, but the cellular mechanisms underlying these differences are not well understood. Using whole‐cell patch‐clamp techniques, we investigated the single‐cell properties and excitatory synaptic features of anatomically identified BCs and AACs in the CA3 region of mouse hippocampal slices. The results showed that BCs had lower threshold for action potential (AP) generation and lower input resistance, narrower AP and afterhyperpolarization than AACs. In addition, BCs fired with higher frequencies and with more modest accommodation compared with AACs. The kinetic properties of excitatory postsynaptic currents (EPSC), the rectification of AMPA receptor‐mediated currents, the fraction of the NMDA receptor‐mediated component in EPSCs, and the EPSC magnitude necessary to evoke an AP were similar in both cell types. However, smaller excitatory postsynaptic potential and lower intensity fiber stimulation in stratum oriens was necessary to drive firing in BCs. Moreover, the rate of spontaneous EPSCs in BCs was higher than in AACs. Neurolucida analysis revealed that the dendrites of BCs in strata radiatum and oriens were longer and more extensively ramified. Since the density of the excitatory synapses was estimated to be comparable in both cell types, we conclude that the more elaborated dendritic arbor of BCs ensures that they receive a larger number of proximal excitatory inputs. Thus, CA3 pyramidal cells more profoundly innervate BCs than AACs, which could explain, at least in part, their distinct spiking behavior under different hippocampal network activities. © 2013 Wiley Periodicals, Inc.     

Different correlations among physiological and morphological properties at single glutamatergic synapses in the rat hippocampus and the cerebellum

Synapses in the mammalian central nervous system show substantial diversity in their physiological and morphological properties. However, the correlations among them have remained elusive. Here, we tried to clarify the correlations by establishing a method to record excitatory postsynaptic currents (EPSCs) at individual synapses and also to observe the morphology at the same time. A pair of pre‐ and postsynaptic neurons were labeled with different fluorescent dyes, and a presynaptic varicosity was selectively stimulated with a θ‐tube glass electrode under conditions in which action potential generation was suppressed. Two representative types of excitatory glutamatergic synapses, one on hippocampal pyramidal neurons and the other on cerebellar Purkinje neurons, were studied. The correlations between the properties of quantal EPSCs (qEPSCs) and those of synaptic morphology were analyzed in rat primary culture preparations. The amplitude and the decay time of qEPSC were correlated with the size of the postsynaptic spine only at hippocampal synapses. In contrast, the size of the presynaptic varicosity was correlated with the size of the postsynaptic spine and the quantal content of evoked EPSCs only at granule neuron–Purkinje neuron synapses in the cerebellum. These results suggest that the interaction between pre‐ and postsynaptic structures and the coupling of postsynaptic responsiveness and the spine morphology differ between cerebellar and hippocampal glutamatergic synapses. Synapse, 2010. © 2010 Wiley‐Liss, Inc.     

Secondhand tobacco smoke exposure differentially alters nucleus tractus solitarius neurons at two different ages in developing non-human primates

Exposing children to secondhand tobacco smoke (SHS) is associated with increased risk for asthma, bronchiolitis and SIDS. The role for changes in the developing CNS contributing to these problems has not been fully explored. We used rhesus macaques to test the hypothesis that SHS exposure during development triggers neuroplastic changes in the nucleus tractus solitarius (NTS), where lung sensory information related to changes in airway and lung function is first integrated. Pregnant monkeys were exposed to filtered air (FA) or SHS for 6 h/day, 5 days/week starting at 50-day gestational age. Mother/infant pairs continued the exposures postnatally to age 3 or 13 months, which may be equivalent to approximately 1 or 4 years of human age, respectively. Whole-cell recordings were made of second-order NTS neurons in transverse brainstem slices. To target the consequences of SHS exposure based on neuronal subgroups, we classified NTS neurons into two phenotypes, rapid-onset spiking (RS) and delayed-onset spiking (DS), and then evaluated intrinsic and synaptic excitabilities in FA-exposed animals. RS neurons showed greater cell excitability especially at age of 3 months while DS neurons received greater amplitudes of excitatory postsynaptic currents (EPSCs). Developmental neuroplasticity such as increases in intrinsic and synaptic excitabilities were detected especially in DS neurons. In 3 month olds, SHS exposure effects were limited to excitatory changes in RS neurons, specifically increases in evoked EPSC amplitudes and increased spiking responses accompanied by shortened action potential width. By 13 months, the continued SHS exposure inhibited DS neuronal activity; decreases in evoked EPSC amplitudes and blunted spiking responses accompanied by prolonged action potential width. The influence of SHS exposure on age-related and phenotype specific changes may be associated with age-specific respiratory problems, for which SHS exposure can increase the risk, such as SIDS and bronchiolitis in infants and asthma in older children.     

The effect of intracellular tetraethylammonium ions on the reversal potential of monosynaptic EPSPs and excitatory amino acids in cultured spinal cord neurones

Summary Using the whole cell patch clamp recording technique, the reversal potential of currents generated by the application of excitatory amino acids and by stimulation of excitatory synapses has been examined in cultured spinal cord neurones, with the partial substitution of intracellular K⁺ by TEA⁺. A previous report (Clements et al. 1986) showed that intracellular injection of TEA into the soma of spinal motoneurones in the cat shifted the reversal of the 1a EPSC to more positive potentials, but the reason for this shift was not clear. The results reported here show that the observed change in the reversal potential for both excitatory amino acid agonists and the excitatory synaptic current, was not due to a change in the relative permeability of the channels to potassium or sodium, but could be explained by a change in driving force for the permeant intracellular ions (K⁺ and/or Cs⁺) on their substitution with the impermeant TEA⁺ ion.     

Properties of inhibitory and excitatory synapses between hippocampal neurons in very low density cultures

The whole cell patch clamp technique was used to examine the electrophysiological properties of embryonic hippocampal neurons maintained in a very low density (VLD) culture prejparation. The goal of these experiments was to establish the viability of the VLD culture as a model system in which to study regulation of neurotransmission at single monosynaptic connections, in the absence of polysynaptic innervation. Depolarization of neurons in the VLD culture revealed voltage-dependent sodium, calcium, and potassium currents which were blocked with, respectively, tetrodotoxin (TTX), cobalt, and tetraethylammonium and 4-aminopyridine. When pairs of neurons were simultaneously recorded, action potentials evoked in presynaptic neurons elicited either excitatory or inhibitory postsynaptic currents (EPSCs or IPSCs, respectively). The dual component “EPSCs” were due to the activation of both types-of postsynaptic, ionotropic glutamate receptors: N-methyl-D-aspartate (NMDA) and non-NMDA receptors. Evoked IPSCs were due to the activation of postsynaptic γ-aminobutyric acid (GABA) receptors. Both excitatory and inhibitory synapses exhibited short term depression in response to high frequency stimulation, although IPSCs were routinely decreased to a much greater degree than EPSCs. Spontaneous miniature EPSCs and IPSCs were found to persist in TTX, were blocked by the same pharmacological antagonists which blocked evoked responses, increased in frequency in response to hyperosmotic solution, and were unaffected by changes in extracellular calcium concentration. mIPSCS were found to occur at a significantly lower frequency than mEPSCs. These experiments indicated that neurotransmission in the VLD cultures occurs in a manner consistent with the quantal hypothesis and, therefore, the VLD culture is a good model for studying excitatory and inhibitory neurotransmission between isolated pairs of neurons. In addition, these experiments, performed under comparable physiological conditions, demonstrated that there are fundamental differences underlying neurotransmitter release between excitatory and inhibitory neurons. © 1994 Wiley-Liss, Inc.     

Role of 5-HT(1) receptor subtypes in the modulation of pain and synaptic transmission in rat spinal superficial dorsal horn

BACKGROUND AND PURPOSE

5-HT receptor agonists have variable nociceptive effects within the spinal cord. While there is some evidence for 5-HT_1A spinally-mediated analgesia, the role of other 5-HT₁ receptor subtypes remains unclear. In the present study, we examined the spinal actions of a range of 5-HT₁ agonists, including sumatriptan, on acute pain, plus their effect on afferent-evoked synaptic transmission onto superficial dorsal horn neurons.

EXPERIMENTAL APPROACH

For in vivo experiments, 5-HT agonists were injected via chronically implanted spinal catheters to examine their effects in acute mechanical and thermal pain assays using a paw pressure analgesymeter and a Hargreave''s device. For in vitro experiments, whole-cell patch-clamp recordings of primary afferent-evoked glutamatergic EPSC were made from lamina II neurons in rat lumbar spinal slices.

KEY RESULTS

Intrathecal (i.t.) delivery of the 5-HT_1A agonist R ± 8-OH-DPAT (30–300 nmol) produced a dose-dependent thermal, but not mechanical, analgesia. Sumatriptan and the 5-HT_1B, 5-HT_1D, 5-HT_1F agonists CP93129, PNU109291 and {"type":"entrez-nucleotide","attrs":{"text":"LY344864","term_id":"1257802930","term_text":"LY344864"}}LY344864 (100 nmol) had no effect on either acute pain assay. R ± 8-OH-DPAT (1 µM) and sumatriptan (3 µM) both reduced the amplitude of the evoked EPSC. In contrast, CP93129, PNU109291 and {"type":"entrez-nucleotide","attrs":{"text":"LY344864","term_id":"1257802930","term_text":"LY344864"}}LY344864 (0.3–3 µM) had no effect on the evoked EPSC. The actions of both R ± 8-OH-DPAT and sumatriptan were abolished by the 5-HT_1A antagonist WAY100635 (3 µM).

CONCLUSIONS AND IMPLICATIONS

These findings indicate that the 5-HT_1A receptor subtype predominantly mediates the acute antinociceptive and cellular actions of 5-HT₁ ligands within the rat superficial dorsal horn.     

Synaptic activity-induced Ca(2+) signaling in avian cochlear nucleus magnocellularis neurons

Neurons of the avian cochlear nucleus magnocellularis (NM) receive glutamatergic inputs from the spiral ganglion cells via the auditory nerve and feedback GABAergic inputs primarily from the superior olivary nucleus. We investigated regulation of Ca²⁺ signaling in NM neurons with ratiometric Ca²⁺ imaging in chicken brain slices. Application of exogenous glutamate or GABA increased the intracellular Ca²⁺ concentration ([Ca²⁺]_i) in NM neurons. Interestingly, GABA-induced Ca²⁺ responses persisted into neuronal maturation, in both standard and energy substrate enriched artificial cerebrospinal fluid. More importantly, we found that electrical stimulation applied to the glutamatergic and GABAergic afferent fibers innervating the NM was able to elicit transient [Ca²⁺]_i increases in NM neurons, and the amplitude of the Ca²⁺ responses increased with increasing frequency and duration of the electrical stimulation. Antagonists for ionotropic glutamate receptors significantly blocked these [Ca²⁺]_i increases, whereas blocking GABA_A receptors did not affect the Ca²⁺ responses, suggesting that synaptically released glutamate but not GABA induced the Ca²⁺ signaling in vitro. Furthermore, activation of GABA_A receptors with exogenous agonists inhibited synaptic activity-induced [Ca²⁺]_i increases in NM neurons, suggesting a role of GABA_A receptors in the regulation of Ca²⁺ homeostasis in the avian cochlear nucleus neurons.     

Cannabinoid CB1 receptors regulate neuronal TNF-α effects in experimental autoimmune encephalomyelitis

Cannabinoid CB1 receptors (CB1Rs) regulate the neurodegenerative damage of experimental autoimmune encephalomyelitis (EAE) and of multiple sclerosis (MS). The mechanism by which CB1R stimulation exerts protective effects is still unclear. Here we show that pharmacological activation of CB1Rs dampens the tumor necrosis factor α (TNFα)-mediated potentiation of striatal spontaneous glutamate-mediated excitatory postsynaptic currents (EPSCs), which is believed to cogently contribute to the inflammation-induced neurodegenerative damage observed in EAE mice. Furthermore, mice lacking CB1Rs showed a more severe clinical course and, in parallel, exacerbated alterations of sEPSC duration after induction of EAE, indicating that endogenous cannabinoids activate CB1Rs and mitigate the synaptotoxic action of TNFα in EAE. Consistently, we found that mice lacking the fatty acid amide hydrolase (FAAH), and thus expressing abnormally high brain levels of the endocannabinoid anandamide, developed a less severe EAE associated with preserved TNFα-induced sEPSC alterations.CB1Rs are important modulators of EAE pathophysiology, and might play a mechanistic role in the neurodegenerative damage of MS patients.     

Loss of striatal cannabinoid CB1 receptor function in attention-deficit / hyperactivity disorder mice with point-mutation of the dopamine transporter

Abnormal dopamine (DA) transmission in the striatum plays a pivotal role in attention-deficit/hyperactivity disorder (ADHD). As striatal DA signalling modulates the endocannabinoid system (ECS), the present study was aimed at investigating cannabinoid CB1 receptor (CB1R) function in a model of ADHD obtained by triple point-mutation in the dopamine transporter (DAT) gene in mice, making them insensitive to cocaine [DAT cocaine-insensitive (DAT-CI) mice]. DAT-CI mice had a marked hyperactive phenotype, and neurophysiological recordings revealed that the sensitivity of CB1Rs controlling GABA-mediated synaptic currents [CB1Rs((GABA)) ] in the striatum was completely lost. In contrast, CB1Rs modulating glutamate transmission [CB1Rs((Glu)) ], and GABA(B) receptors were not affected in this model of ADHD. In DAT-CI mice, the blockade of CB1R((GABA)) function was complete even after cocaine or environmental manipulations activating the endogenous DA-dependent reward system, which are known to sensitize these receptors in control animals. Conversely, the hedonic property of sucrose was intact in DAT-CI mice, indicating normal sweet perception in these animals. Our results point to CB1Rs as novel molecular players in ADHD, and suggest that therapeutic strategies aimed at interfering with the ECS might prove effective in this disorder.