Unit study in cat claustrum of the effects of iontophoretic neurotransmitters and correlations with the effects of activation of some afferent pathways

Glutamate (Glut), acetylcholine (ACh) and dopamine (DA) were iontophoretically applied on cat claustral neurons. Glut did not affect all the neurons; ACh had both excitatory and inhibitory effects, while DA was prevalently inhibitory. An analysis was made of the time-course of excitatory and inhibitory responses on the basis of the mean firing rate variations during and after ACh and DA release. Three types of responses are described for each drug: short lasting inhibition, long lasting inhibition and long lasting excitation. The experimental data were statistically elaborated. The effects of ACh and of DA were compared with those of activation obtained by sensorial peripheric and thalamic stimulations. ACh could be supposed to be the transmitter of most of the inhibitory terminals of these sensitive afferences to the claustrum.     

Projections from the claustrum to the occipital cortex in the guinea pig

The existence of ipsilateral claustrocortical projection to the occipital area Oc 2.2 (Wree et al. 1981) was demonstrated in four guinea pigs. The neurons labelled by retrogradely transported HRP were localized in the caudal half of the claustrum dorsale (in its dorsomedial and central part); small and medium-sized oval and multipolar neurons preponderated.     

Electrophysiological and microiontophoretic studies with buspirone: influence on the firing rate of central monoaminergic neurons and their responsiveness to dopamine, clonidine or GABA

The influence of an i.v. perfusion of buspirone on the firing rate of central monoaminergic neurons was studied in rats anaesthetized with chloral hydrate. Buspirone increased the firing rate of A10 dopaminergic neurons and blocked the inhibitory effect of iontophoretically applied dopamine on these neurons. A slight attenuation of the inhibitory effect of iontophoretically applied GABA was also observed. Buspirone increased the firing rate of locus coeruleus (LC) noradrenergic neurons and induced an attenuation of the inhibitory effect of iontophoretically applied clonidine. A slight attenuation of the inhibitory effect of iontophoretically applied GABA was also observed. Furthermore buspirone was a very potent inhibitor of the firing rate of dorsal raphe (DR) serotonergic neurons. It is concluded that activation of A10 neurons by buspirone is due to blockade of dopaminergic autoreceptors and that activation of LC neurons is related to blockade of alpha-2 autoreceptors. The significance of the interaction with gabaergic inhibition is unclear. The mechanisms involved in the inhibition of DR neurons remain to be investigated.     

Neuroprotective role of delta-opioid receptors in cortical neurons

We recently demonstrated that delta-opioid receptor (DOR) activation protects cortical neurons against glutamate-induced injury. Because glutamate is a mediator of hypoxic injury in neurons, we hypothesized that DOR is involved in neuroprotection during O2 deprivation and that its activation/inhibition may alter neuronal susceptibility to hypoxic stress. In this work, we tested the effect of opioid receptor activation and inhibition on cultured cortical neurons in hypoxia (1% O2). Cell injury was assessed by lactate dehydrogenase release, morphology-based quantification, and live/dead staining. Our results show that 1) immature neurons (days 4 and 6) were not significantly injured by hypoxia until 72 h of exposure, whereas day 8 neurons were injured after only 24-h hypoxia; 2) DOR inhibition (naltrindole) caused neuronal injury in both day 4 and day 8 normoxic cultures and further augmented hypoxic injury in these neurons; 3) DOR activation ([D-Ala2,D-Leu5]enkephalin) reduced neuronal injury in day 8 cultures after 24 h of normoxic or hypoxic exposure and attenuated naltrindole-induced injury with prolonged exposure; and 4) mu- or kappa-opioid receptor inhibition (beta-funaltrexamine or nor-binaltorphimine) had little effect on neurons in either normoxic or hypoxic conditions. Collectively, these data suggest that DOR plays a crucial role in neuroprotection in normoxic and hypoxic environments.     

尾核阿片μ受体参与电针及皮层SmⅠ区对束旁核伤害性反应的抑制

为探索尾核（ｃａｕｄａｔｅｎｕｃｌｅｕｓ,Ｃｄ）是否参与电针及皮层体感运动Ⅰ区（ｓｅｎｓｏｒｉｍｏｔｏｒａｒｅａⅠｏｆｔｈｅｃｅｒｅｂｒａｌｃｏｒｔｅｘ,ＳｍⅠ）对束旁核（ｐａｒａｆａｓｃｉｃｕｌａｒｎｕｃｌｅｕｓ,Ｐｆ）神经元伤害性反应的调节,以及Ｃｄ中阿片受体是否参与并通过何种受体参与这一调节,本实验用Ｃｄ头部化学毁损及微量注射阿片受体拮抗剂的方法,观察到Ｃｄ毁损前电针及兴奋皮层均可抑制Ｐｆ的伤害性反应,而毁损后这种抑制效应消失;注射纳洛酮或阿片μ受体拮抗剂βＦＮＡ后,电针及兴奋皮层ＳｍⅠ区对Ｐｆ伤害性反应的抑制作用被取消,而分别注射δ和κ受体拮抗剂ＩＣＩ１７４,８６４和ｎｏｒＢＮＩ则不产生影响。基于已证明大脑皮层参与电针对Ｐｆ伤害性反应的调节,本结果提示：Ｃｄ参与针刺镇痛中皮层ＳｍⅠ区对Ｐｆ神经元伤害性反应的抑制,Ｃｄ中阿片肽主要通过μ受体参与抑制作用。     

Light and electron-microscopic study of leucine enkephalin immunoreactivity in the cat claustrum

The claustrum is a complex telencephalic structure owing to its reciprocal connectivity with most—if not all—cortical areas. However, there is a paucity of data in the literature concerning its histochemical components, including opioid peptide neurotransmitters. The aim of the present study was to examine the morphology, distribution and ultrastructure of leucine-enkephalin-immunoreactive (Leu-enk-ir) neurons and fibers in the dorsal claustrum (DC) of the cat. Seven healthy, adult male and female cats were used in our study. All animals received humane care. They were irreversibly anesthetized and transcardially perfused with fixative. Brains were removed, postfixed, blocked and sectioned. Sections were incubated with polyclonal anti-Leu-enk antibodies using the Avidin–Biotin–Peroxidase Complex method. Leu-enk-ir neurons and fibers were distributed throughout the DC. Some of the neurons were lightly-stained, while others were darkly-stained. Light-microscopically, they varied in shape: oval, fusiform, multipolar and irregular. With regard to size, they were categorized as small (15?μm or less in diameter), medium (16–20?μm in diameter) and large (21?μm or more in diameter). No specific pattern of regional distribution was found. On the electron microscope level, immunoproduct was observed in neurons, dendrites and terminal boutons. Different types of Leu-enk-ir neurons differ in their ultrastructural features, including two types of synaptic boutons. No gender-specific features were observed. In conclusion, it is our hope that our study will serve to contribute to a better understanding of the functional neuroanatomy of the DC in the cat, and that it can be extrapolated and applied to other mammals, including humans.     

Non-adrenergic mechanisms of inhibition of the contractile activity of the cat small intestine by histamine, bradykinin, and met-enkephalin

I. a. histamine and bradykinin caused both an activation and an inhibition of jejunal contractile activity. The inhibitory effect was preserved after blockade of muscarinic cholinoreceptors, alpha- and beta-adrenoceptors, and abolished with the nicotinic cholinoceptor blockade. Metenkephalin inhibited the jejunal contractile activity after first activating it. The inhibitory effect of the peptide was preserved after blockade of alpha- and beta-adrenoceptors as well as the nicotinic cholinoceptors. The data obtained suggest that the non-adrenergic inhibitory effect of metenkephalin on intestinal contractions was the result of its depressing action on motor cholinergic neurons, whereas the inhibition of intestinal contractile activity with histamine and bradykinin resulted from their activating action on cholinergic interneurons which activate non-adrenergic inhibitory neurons through nicotinic cholinoceptors.     

Ischemia induces short- and long-term remodeling of synaptic activity in the hippocampus

One of the most vulnerable areas to ischemia or hypoglycemia is CA1 hippocampal region due to pyramidal neurons death. Glutamate receptors are involved together with protein-kinase C and nitric oxide synthase. Long-term potentiation (LTP) is generated in anoxic or hypoglycemic conditions via activation of NMDA while inhibition of these receptors atenuates this response. Protein-kinase C and nitric oxide synthase are involved in anoxic LTP mechanism. Postischemic neurons are hyperexcitable in CA3 area while CA1 pyramidal neurons degenerate and dissapear. Changes of glutamate receptors triggered by ischemia and hypoglycemia are discussed in this review.     

Spontaneous electrical activity of the claustrum

The dynamics of spontaneous electrical activity of the rostral, dorsal, ventral, and caudal parts of the claustrum were studied in chronic experiments on 12 dogs with implanted electrodes during extinction of orienting reactions and during biologically meaningful changes in the animals' functional state. In the animal at rest, electrical activity of the claustrum was similar to that of subcortical structures, but in a state of attention, its activity began to resemble that of the electrocorticogram, in agreement with the view that this nucleus occupies an intermediate position between cortical and subcortical structures. Enhancement of fast activity in the claustrum with the appearance of orienting reactions was expressed more intensively, and during extinction it was preserved for a longer time, in the dorsal and ventral parts of the claustrum than in the rostral and caudal parts, evidence of structural heterogeneity of the nucleus. Under conditions of increased food excitability the amplitude and frequency of the fast waves recorded from the rostral and caudal parts of the claustrum were selectively increased, whereas during reflexes to a rejected stimulus (acids) and in defensive reflexes they were increased in recordings from the dorsal and ventral parts, on which basis the existence of specialized efferent zones participating in biologically different types of activity can be postulated. Enhancement of fast activity in the claustrum during reflexes to food and acid differed in its genesis: The switch from the first to the second took place through a phase of suppression of the first. Differentiation of a food-conditioned stimulus was accompanied by the development of active inhibition in the "food zones" of the claustrum and by enhancement of activity in the "zones of orienting reactions."A. A. Bogomolets Institute of Physiology, Academy of Sciences of the Ukrainian SSR, Kiev. Translated from Neirofiziologiya, Vol. 12, No. 2, pp. 155–164, March–April, 1980.     

Application of proteasomal inhibitors to mouse sympathetic neurons activates the intrinsic apoptotic pathway

Proteasomal dysfunction may play a role in a number of neurodegenerative conditions, and in particular Parkinson's disease (PD) and related Lewy body (LB) diseases. Application of proteasomal inhibitors to neuronal cell culture systems is associated with survival-promoting effects or with cell death depending on the model system. We have applied pharmacological proteasomal inhibitors to cultured neonatal mouse sympathetic neurons in order to investigate whether these catecholaminergic neurons, which are affected in PD, are sensitive to proteasomal inhibition and, if so, which cell death pathway is activated. We report here that proteasomal inhibition leads to apoptotic death of mouse sympathetic neurons. This death is accompanied by caspase 3 activation and cytochrome c release from the mitochondria and is abrogated by caspase inhibition. Bax deletion prevented both cytochrome c release and caspase 3 activation, and also provided complete protection against proteasomal inhibition-induced death. Bcl-2 overexpression achieved a similar survival-promoting effect. There was no change in Bax levels following proteasomal inhibition, suggesting that Bax itself is not regulated by the proteasome in this cell culture system, and that a primary increase in Bax is unlikely to account for death. In contrast, levels of the BH3-only protein, Bim, increased with proteasomal inhibition. We conclude that proteasomal inhibition of mouse sympathetic neurons activates the intrinsic apoptotic pathway involving bcl-2 family members and the mitochondria.     

Dopamine sensitive neurons of the arcuate region of the hypothalamus and their role in regulating the gonadotropic function of the pituitary]

The effect of the microiontophoretic application of dopamine (DA) into the arcuate region of the hypothalamus on the sensitivity of single neurons and on the plasma and hypophysis LH levels was examined at various stages of the estrous cycle. During the estrous cycle in rats there was no significant differences in the relative number of neurons showing activation and inhibition or non-responsive to DA. However, in the first half of proestrus (P) a significant increase in the number of neurons with the excitative reaction to the iontophoretic application of DA was observed. At all the stages of the cycle, except the second half of P, the excitative reaction of neurons correlated with increased LH level in the plasma. In the hypophysis only in diestrus-2 there was a significant increase of the LH level in response to the iontophoretic application of DA in the arcuate neurons of the hypothalamus.     

Topography of Gng2- and NetrinG2-Expression Suggests an Insular Origin of the Human Claustrum

The claustrum has been described in the forebrain of all mammals studied so far. It has been suggested that the claustrum plays a role in the integration of multisensory information: however, its detailed structure and function remain enigmatic. The human claustrum is a thin, irregular, sheet of grey matter located between the inner surface of the insular cortex and the outer surface of the putamen. Recently, the G-protein gamma2 subunit (Gng2) was proposed as a specific claustrum marker in the rat, and used to better delineate its anatomical boundaries and connections. Additional claustral markers proposed in mammals include Netrin-G2 in the monkey and latexin in the cat. Here we report the expression and distribution of Gng2 and Netrin-G2 in human post-mortem samples of the claustrum and adjacent structures. Gng2 immunoreactivity was detected in the neuropil of the claustrum and of the insular cortex but not in the putamen. A faint labelling was present also in the external and extreme capsules. Double-labelling experiments indicate that Gng2 is also expressed in glial cells. Netrin-G2 labelling was seen in neuronal cell bodies throughout the claustrum and the insular cortex but not in the medially adjacent putamen. No latexin immunoreactive element was detected in the claustrum or adjacent structures. Our results confirm that both the Gng2 and the Netrin-G2 proteins show an affinity to the claustrum and related formations also in the human brain. The presence of Gng2 and Netrin-G2 immunoreactive elements in the insular cortex, but not in the putamen, suggests a possible common ontogeny of the claustrum and insula.     

Microinjections of heat shock protein 70 kDa into the nucleus reticularis pontis oralis induce inhibition of rapid eye movement sleep in pigeons

Recently it was indicated that microinjections of heat shock proteins 70 kDa (Hsp70) into the third ventricle of brain in pigeons results in an increase in the duration of slow wave sleep and a decrease in somato-visceral indices. It is suggested that Hsp70 effect may be related to GABA(A) receptors activation in the preoptic area of the hypothalamus. However, what transmitter mechanisms of activation are related to the removal effect (in 2-3 hrs) of rapid eye movement sleep inhibition still remains poorly understood. To solve this problem in the present study, microinjections of Hsp70 into the Nucleus reticularis pontis oralis (NRPO) were done. It is well known that cholinergic neurons of the NRPO are crucial for rapid eye movement sleep generation. The data show that Hsp70 produces more early (for first two hrs) a decrease in number of episodes and total time of rapid eye movement sleep, a diminution of electroencephalogram (EEG) power spectra in the 9-14 Hz band, a decrease in contractile muscle activity and brain temperature. It is suggested that Hsp70 effects are realized due to activation of GABA(A) receptors in the NRPO and induced inhibition of cholinergic mechanisms of rapid eye movement sleep triggering. The microinjections of Hsp70 into the NRPO increase the slow wave sleep total time with long latency (for 8-12 hrs). This effect may be related to influence of Hsp70 on neurons population, which are responsible for slow wave sleep maintenance outside the NRPO.     

Inhibition of locus coeruleus neuronal activity by beta-phenylethylamine

The effect of beta-phenylethylamine (PEA) on brain noradrenaline (NA) neurons in the rat locus coeruleus (LC) was analyzed using single unit recording techniques including microiontophoretic methodology. Systemic injection of low doses of PEA consistently produced an instantaneous and dose-dependent inhibition of firing rate of the LC neurons. The effect was strongly antagonized by administration of the alpha 2-receptor antagonist yohimbine (1 mg/kg, i.v.) or by depletion of endogenous stores of NA by pretreatment with reserpine (10 mg/kg, i.p., 6 h), but unaffected by inhibition of tyrosine hydroxylase (alpha-met-hyl-p-tyrosine (alpha-MT), 250 mg/kg, i.p., 30 min). In contrast, the inhibitory effect of PEA on the LC neurons was strongly potentiated by pretreatment with the selective monoamine oxidase (MAO) - B inhibitor pargyline (2 mg/kg, i.p., 1 h), but, unexpectedly, also by pretreatment with the MAO-A selective inhibitors clorgyline (2 mg/kg, i.p., 1 h) or FLA 336 (2 mg/kg, i.p., 1 h). When microiontophoretically applied directly onto the LC neurons, PEA produced inhibition of a majority of the NA neurons. This action was prevented by intravenous injection of yohimbine (2.5 mg/kg). The results suggests that the action of PEA on NA neurons in the LC is an indirect effect, requiring availability of a reserpine-sensitive storage pool of NA, and mediated via activation of central alpha 2-receptors within the LC.     

The claustrum in the dog brain.

The structure of the claustrum was studied in the dog brain using Weigert's, Klüver-Barrera's, Manns' and Nissl's methods. It consists of two main parts arranged one above the other. The dorsal part is situated in the depth of the neocortex and extends from the gyrus orbitalis to the gyrus compositus posterior. The ventral part of the claustrum underlying the olfactory cortex continues from the cuadal fragments of the olfactory peduncle to the entorhinal area, where it fuses with its deep layers. The claustrum can be regarded as a fragment of a bigger cellular formation present in different parts of the hemisphere.     

Cholinergic inhibition of electrogenic sodium absorption in the guinea pig distal colon

Submucosal cholinergic and noncholinergic neurons in intestines have been shown to be involved in regulating epithelial transport functions, particularly stimulating Cl(-) secretion. This study investigates the role of submucosal cholinergic neurons in regulating electrogenic Na(+) absorption in distal colon. Amiloride-sensitive short-circuit current (I(sc)) and (22)Na(+) flux were measured in mucosal and mucosal-submucosal preparations mounted in Ussing chambers. In the mucosal preparation, carbachol (CCh) added to the serosal side inhibited amiloride-sensitive I(sc) and amiloride-sensitive (22)Na(+) absorption. The inhibitory effect of CCh was observed at approximately 0.1 microM, and maximum inhibition of approximately 70% was attained at approximately 30 microM (IC(50) = approximately 1 microM). CCh-induced inhibition of amiloride-sensitive I(sc) was almost totally abolished by 10 microM atropine. Treatment of the tissue with ionomycin markedly reduced amiloride-sensitive I(sc), but a subsequent addition of CCh further decreased it. Also, CCh still had an inhibitory effect, although significantly attenuated, after the tissue had been incubated with a low-Ca(2+) solution containing ionomycin and BAPTA-AM. Applying electrical field stimulation to submucosal neurons in the mucosal-submucosal preparation resulted in inhibition of amiloride-sensitive I(sc), approximately 33% of this inhibition being atropine sensitive. Physostigmine inhibited amiloride-sensitive I(sc), this effect being abolished by atropine. In conclusion, submucosal cholinergic and noncholinergic neurons were involved in inhibiting electrogenic Na(+) absorption in colon. This inhibition by cholinergic neurons was mediated by muscarinic receptor activation.     

Regulation of Hypothalamo-Pituitary-Adrenocortical Responses to Stressors by the Nucleus of the Solitary Tract/Dorsal Vagal Complex

Hindbrain neurons in the nucleus of the solitary tract (NTS) are critical for regulation of hypothalamo-pituitary-adrenocortical (HPA) responses to stress. It is well known that noradrenergic (as well as adrenergic) neurons in the NTS send direct projections to hypophysiotropic corticotropin-releasing hormone (CRH) neurons and control activation of HPA axis responses to acute systemic (but not psychogenic) stressors. Norepinephrine (NE) signaling via alpha1 receptors is primarily excitatory, working either directly on CRH neurons or through presynaptic activation of glutamate release. However, there is also evidence for NE inhibition of CRH neurons (possibly via beta receptors), an effect that may occur at higher levels of stimulation, suggesting that NE effects on the HPA axis may be context-dependent. Lesions of ascending NE inputs to the paraventricular nucleus attenuate stress-induced ACTH but not corticosterone release after chronic stress, indicating reduction in central HPA drive and increased adrenal sensitivity. Non-catecholaminergic NTS glucagon-like peptide 1/glutamate neurons play a broader role in stress regulation, being important in HPA activation to both systemic and psychogenic stressors as well as HPA axis sensitization under conditions of chronic stress. Overall, the data highlight the importance of the NTS as a key regulatory node for coordination of acute and chronic stress.     

Involvement of CRF on the anorexic effect of GLP-1 in layer chicks

Glucagon-like peptide-1 (GLP-1) is recognized as an anorexic peptide in the brain of chicks. However, the mechanism underlying the inhibition of feeding has not been well studied. It is reported that GLP-1 activates neurons containing corticotrophin-releasing factor (CRF) in the brain of mammals. Since CRF is also an anorexic peptide, it is possible that the anorexic effect of GLP-1 is mediated by CRF in chicks. The present study was carried out to test this. First, we determined plasma corticosterone (CORT) concentrations after intracerebroventricular (ICV) injection of GLP-1 and found that this treatment increased CORT release in layer chicks. The CORT-releasing effect was partly attenuated by co-injection of astressin, a CRF receptor antagonist, demonstrating that GLP-1 stimulated CORT secretion by activation of CRF neurons. CRF neurons also appear to be involved in mediating the inhibition of food intake by GLP-1 because this effect was also partly attenuated by astressin. Furthermore, we demonstrated that the anorexic effect of GLP-1 was weaker in broiler than layer chicks. The present results suggest that the anorexic effect of GLP-1 might be mediated by CRF neurons in the chick brain and that the sensitivity of the inhibitory response to GLP-1 differs between chick strains.     

Hypertension alters GABA receptor-mediated inhibition of neurons in the nucleus of the solitary tract

Previous studies have demonstrated that microinjection of baclofen, a GABA(B) receptor agonist, into the nucleus of the solitary tract (NTS) results in an enhanced pressor response in hypertensive (HT) rats compared with normotensive (NT) rats, suggesting a possible alteration in the responses of neurons in this area to activation of GABA(B) receptors. The following studies were designed to determine whether HT alters the sensitivity of neurons in the NTS to GABA receptor agonists. Sham-operated NT and unilateral nephrectomized, renal-wrap HT Sprague-Dawley rats were anesthetized, and the responses of NTS neurons receiving aortic nerve (AN) afferent inputs to iontophoretic application of GABA, the GABA(A) receptor agonist muscimol, and the GABA(B) agonist baclofen were examined. The AN input was classified as monosynaptic (MSN) if the cell responded to each of two stimuli separated by 5 ms with an action potential. If the cell did not respond, the input was considered polysynaptic (PSN). In MSNs, inhibition of AN-evoked discharge by GABA was not altered in 1 wk of HT but was reduced in 4 wk of HT, whereas in PSNs, sensitivity to GABA was reduced at 1 and 4 wk of HT. In HT rats, inhibition of AN-evoked discharge by baclofen was enhanced in MSNs, but not in PSNs, after 1 and 4 wk of HT, whereas inhibition by muscimol was reduced in MSNs and PSNs at 1 and 4 wk of HT. Changes in sensitivity to muscimol and baclofen within MSNs were the same whether the MSN received a slowly or a rapidly conducted AN afferent input. The results demonstrate that early in HT the sensitivity of NTS neurons to inhibitory amino acids is altered and that these changes are maintained for > or =4 wk. The alterations are dependent on the subtype of GABA receptor being activated and whether the neuron receives a mono- or polysynaptic baroreceptor afferent input.