Calcium antagonist, adenosine A1, and muscarinic bindings in rat hippocampus after transient ischemia

The protective roles of Ca2+ channel blockers against ischemic hippocampal damage are still debated. We used autoradiography to study postischemic L-type Ca2+ channels (1,4-dihydropyridine Ca2+ channel blocker binding), adenosine A1 receptors, and muscarinic cholinergic receptors in the rat hippocampus using [3H]PN200-110 (PN), [3H]cyclohexyladenosine (CHA), and [3H]quinuclidinyl benzilate (QNB), respectively, in 49 rats subjected to 20 minutes of forebrain ischemia. The rats were decapitated after 1 (n = 7), 3 (n = 7), 6 (n = 8), 12 (n = 7), 24 (n = 6), 48 (n = 6), or 168 (n = 8) hours of recirculation; eight control rats were sham-operated but experienced no cerebral ischemia. Reduced receptor binding preceding the delayed death of CA1 pyramidal cells was first observed in the stratum oriens of the CA1 subfield. Significant reductions in [3H]PN, [3H]CHA, and [3H]QNB bindings of this stratum compared with control were noticed after 3 (35%, p less than 0.01), 12 (31%, p less than 0.01), and 1 (10%, p less than 0.05) hours of recirculation, respectively. By 168 hours after ischemia (when the populations of CA1 pyramidal cells were depleted) all strata in the CA1 subfield had lost most of their receptor sites, and [3H]PN, [3H]CHA, and [3H]QNB bindings in the stratum oriens were decreased to 23%, 30%, and 63% of control (p less than 0.01). Although [3H]PN binding in the CA3 subfield did not change significantly during 168 hours after ischemia, the histologically intact dentate gyrus exhibited a 31% loss of binding sites compared with control (p less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)     

Postischemic alteration of muscarinic acetylcholine, adenosine A1 and calcium antagonist binding sites in selectively vulnerable areas: an autoradiographic study of gerbil brain.

We performed receptor autoradiography to determine sequential alterations in the binding of muscarinic cholinergic and adenosine A1 receptors and of a voltage dependent L-type calcium channel blocker 1 h-1 month after transient cerebral ischemia in the gerbil brain. [3H]Quinuclidinyl benzilate (QNB), [3H]cyclohexyladenosine (CHA) and [3H]PN200-110 were used to label muscarinic and adenosine A1 receptors and L-type calcium channels, respectively. Transient ischemia was induced for 10 min. [3H]QNB and [3H]CHA binding showed no significant alteration in selectively vulnerable areas at an early stage (1-24 h) of recirculation. However, the dentate molecular layer which was resistant to ischemia revealed a significant decrease in the [3H]CHA binding sites 24 h after ischemia. Thereafter, the [3H]QNB and [3H]CHA binding showed significant reduction in most of selectively vulnerable areas. Marked reduction was especially found in the dorsolateral part of striatum and the hippocampal CA1 sector which was the most vulnerable to ischemia. In contrast, [3H]PN200-110 binding showed a transient elevation in the hippocampal CA1 sector, the dentate molecular layer and the thalamus 1 h of recirculation. However, the striatum and neocortex revealed no alteration in the [3H]PN200-110 binding. Thereafter, the reduction in the [3H]PN200-110 binding was seen only in the dorsolateral part of the striatum and the hippocampal CA1 sector. The results suggest that transient cerebral ischemia can cause the alterations in the binding of muscarinic cholinergic and adenosine A1 receptors and of L-type calcium channel blocker in most of selectively vulnerable areas.(ABSTRACT TRUNCATED AT 250 WORDS)     

Autoradiographic analysis of second-messenger systems in the gerbil hippocampus following repeated brief ischemic insults.

Using [3H]inositol 1,4,5-triphosphate (IP3), [3H]phorbol 12,13-dibutyrate (PDBu) and [3H]forskolin, we performed quantitative autoradiography to determine sequential alterations in second-messenger systems in the gerbil hippocampus following repeated brief ischemic insults. Changes following three 2-min ischemic insults were compared with those following single 2- or 6-min ischemia. [3H]IP3 binding was extremely sensitive to ischemic insult, and more than 80% of the binding sites were lost after destruction of CA1 pyramidal cells following 6-min ischemia and three 2-min ischemic insults. Furthermore, a 30% reduction was observed after 2-min ischemia which leads to no neuronal loss. [3H]PDBu binding in the CA1 subfield decreased by 1 day after three 2-min ischemic insults and by 4 days after 6-min ischemia, and 40-50% reductions were observed at 1 month. In contrast, [3H]forskolin binding was relatively preserved. [3H]PDBu and [3H]forskolin binding transiently increased early in the reperfusion period. We also observed a difference in the pattern and severity of alterations between repeated ischemic insults and single ischemia.     

Preserved neurotransmitter receptor binding following ischemia in preconditioned gerbil brain

Preconditioning the brain with sublethal ischemia induces tolerance to subsequent ischemic insult. Using [3H]quinuclidinyl benzilate (QNB), [3H]MK 801, [3H]cyclohexyladenosine, [3H]muscimol, and [3H]PN200-110, we investigated the alterations in neurotransmitter receptor and calcium channel binding in the gerbil hippocampus following ischemia with or without preconditioning. Two-minute forebrain ischemia, which produced no neuronal damage, resulted in no alterations in binding except for a slight reduction in [3H]QNB binding in the CA1 subfield. Three-minute ischemia destroyed the majority of CA1 pyramidal cells and caused, in CA1, reductions in binding of all ligands used. Preconditioning with 2-min ischemia followed by 4 days of reperfusion protected against CA1 neuronal damage and prevented the reductions in binding although [3H]QNB and [3H]PN200-110 binding transiently decreased in the early reperfusion period, suggesting down-regulation. Thus, preconditioning protects against damage to the neurotransmission system as well as histopathological neuronal death.     

Autoradiographic analysis of second messenger and neurotransmitter system receptors in the gerbil hippocampus following transient forebrain ischemia.

Changes in second messenger and neurotransmitter system receptor ligand binding induced by transient forebrain ischemia were studied in the gerbil hippocampus. The animals were allowed variable periods of recovery ranging from 2 h to 7 days after 5-min bilateral carotid artery occlusion. The binding of second messenger systems ([3H]inositol 1,4,5-trisphosphate ([3H]IP3)to inositol 1,4,5-triphosphate, [3H]forskolin to adenylate cyclase and [3H]phorbol 12,13-dibutylate to protein kinase C) and neurotransmitter receptor systems ([3H]PN200-110 to L-type calcium channels. [3H]N6-cyclohexyl-adenosine to adenosine A1 and [3H]quinuclidinyl benzilate to muscarinic cholinergic receptor) were assayed using quantitative autoradiography. In the CA1 subfield, 2 h after ischemia, [3H]IP3, [3H]forskolin, and [3H]quinuclidinyl benzilate binding activities significantly decreased by 25, 17 and 13%, respectively, though no morphological abnormalities were obvious. Six hours after ischemia, the [3H]phorbol 12,13-dibutylate binding activity in the stratum oriens of the CA1 subfield increased by 15%. One day after ischemia, [3H]PN200-110 binding activity in this subfield decreased by 26%, and 7 days after ischemia, [3H]phorbol 12,13-dibutylate and [3H]N6-cyclohexyl-adenosine receptor binding activities decreased in this subfield. In particular, at 7 days after ischemia, [3H]IP3 binding activity in the CA1 subfield showed a complete decline. In the CA3 subfield, [3H]PN200-110 binding activity decreased 2 days after ischemia, and [3H]IP3 and [3H]N6-cyclohexyl-adenosine binding activities decreased 7 days after ischemia. In the dentate gyrus, the structure of which remained histologically intact after ischemic insult, [3H]IP3 and [3H]forskolin binding activities decreased 7 days after ischemia. In contrast, the [3H]phorbol 12,13-dibutylate binding activity increased in the molecular layer of the dentate gyrus 7 days after ischemia. These results indicate that marked alteration of intracellular signal transduction precedes neuronal damage in the hippocampal CA1 subfield and that the histologically intact CA3 and dentate gyrus also shows modulated neuronal transmission after ischemia.     

Regional changes in [H]inositol 1,4,5-triphosphate binding in the gerbil brain following repeated sublethal ischemia

We investigated the regional changes in [³H]inositol 1,4,5-triphosphate (IP₃) binding in the brain following ischemia using in vitro autoradiography. Three 2-min ischemic insults at 1-hr intervals and a 6-min period of ischemia were induced in gerbils and they were killed after 1, 4, and 28 days. Normal animals had high [³H]IP₃ binding in the CA1 subfield of the hippocampus and the striatum. The binding in the CA1 decreased strikingly after both 6-min ischemia and three 2-min ischemic insults. The [³H]IP₃ binding also decreased in the lateral striatum after three 2-min ischemic insults but not after 6 min of ischemia. Histological observations confirmed neuronal damage to these areas of reduced binding. By contrast, we found a marked increase in [³H]IP₃ binding in the ventral thalamus 28 days after three 2-min ischemic insults. Histological observations with Nissl staining revealed an accumulation of fine granular deposits there. Thus, repeated ischemic insults produced more extensive neuronal damage and changes in [³H]IP₃ binding than a single equivalent period of ischemia. The increased [³H]IP₃ binding in the thalamus coincidentally with an accumulation of Nissl-positive granules at the chronic stage after repeated ischemia is of considerable interest.     

Quantitative autoradiographic analysis of muscarinic cholinergic and adenosine A1 binding sites after transient forebrain ischemia in the gerbil

The influence of transient forebrain ischemia on adenosine A₁ and muscarinic cholinergic receptors in the gerbil brain 1–27 days after recirculation was studied. The topographical distribution and the alteration in the adenosine A₁ and muscarinic receptor sites were analyzed by means of quantitative receptor autoradiography using [³H]cyclohexyladenosine ([³H]CHA) and [³H]quinuclidinyl benzilate ([³H]QNB), respectively. In most regions examined, the temporal profiles of the alteration of the receptor density were in accordance with the histopathological findings. [³H]CHA binding activity decreased suddenly after neuronal damage, while [³H]QNB grain density showed a gradual decrease in the dorsolateral caudate-putamen and in the CA₁ subfield of the hippocampus. In the caudate-putamen, [³H]CHA and [³H]QNB binding activity in the dorsal aspect was markedly reduced 1–27 days after ischemia. [³H]CHA binding activity in the ventromedial region of the caudate-putamen also decreased 1–3 days after ischemia, though neuronal damage was restricted to the dorsolateral aspect. Neuronal death in CA₁ was preceded by the decrease in [³H]QNB binding activity in the stratum radiatum 1 and 2 days after ischemia. Marked decrease in [³H]QNB and [³H]CHA binding activity was noted in the CA₁ subfield 3–27 days after recirculation. Three to 27 days after ischemia, the A₁ binding activities in the CA₃ subfield of the hippocampus and int he dentate gyrus were reduced despite the normal appearance of these areas throughout the reperfusion period. Muscarinic binding sites in the CA₃ subfield were also reduced 27 days after ischemia. Despite minimal neuronal damage in the lateral septal nucleus and in the substantia nigra, the A₁ binding activity in these regions was reduced by 70% and 50%, respectively. These results provide further evidence that the muscarinic receptors in the dorsolateral region of the caudate-putamen are localized postsynaptically on small and medium-sized neurons and that those in the CA₁ subfield of the hippocampus are localized on the CA₁ pyramidal cells.     

Autoradiographic visualization of adenosine A1 receptors in the gerbil hippocampus: changes in the receptor density after transient ischemia

N6-cyclohexyl-[3H]adenosine [( 3H]CHA) was used for the in vitro visualization of the hippocampal adenosine A1 receptors in the gerbil. The strata radiatum and oriens of the hippocampus showed particularly high binding activity. Depletion of pyramidal cells and consequent severe decrease in [3H]CHA binding activity in the CA1 subfield were observed after transient ischemic insult. These results suggest that most adenosine receptors in the CA1 region are localized in association with pyramidal cells.     

Induction of tolerance to ischemia: Alterations in second-messenger systems in the gerbil hippocampus

Preconditioning the brain with sublethal ischemia protects against neuronal damage following subsequent ischemic insult. Using [3H]inositol 1,4,5-triphosphate (IP3), [3H]phorbol 12,13-dibutyrate (PDBu), [3H]cyclic adenosine monophosphate (cAMP) and [3H]rolipram, we performed quantitative autoradiography to determine postischemic alterations in second-messenger systems in the gerbil hippocampus following preconditioning the brain with sublethal ischemia. At 7 days of reperfusion, no alterations were observed in brains subjected to 2 min of forebrain ischemia which produced no neuronal damage. However, 3-min ischemia caused a 75% reduction in [3H]IP3 binding (p < 0.01 vs. control) and 15-25% reductions in [3H]forskolin (p < 0.01 vs. control), [3H]cAMP (p < 0.05 vs. control), and [3H]rolipram (p < 0.01 vs. control) binding in the CA1 subfield coincident with histopathological CA1 pyramidal cell destruction, but no significant alterations in [3H]PDBu binding. Preconditioning the brain with 2 min of ischemia followed by 4 days of reperfusion prevented both histopathological cell death and the reductions in binding following subsequent 3 min of ischemia. Interestingly, [3H]IP3 and [3H]rolipram binding in CA1 showed a transient reduction, by 30% and 20% (both p < 0.01 vs. control), respectively, in the early reperfusion period. This downregulation of the IP3 system may play a role in the protection against cell death.     

Regional variations in particulate cyclic AMP dependent-protein kinase binding activity in the gerbil hippocampus following transient forebrain ischemia by [3H]cyclic AMP binding.

Changes in the binding of [3H]cyclic AMP as an indicator of particulate cyclic AMP-dependent protein kinase (AMP-DPK) binding activity following transient forebrain ischemia were studied in the gerbil using in vitro autoradiography. [3H]Cyclic AMP binding in the strata pyramidale and lacunosum-moleculare of the hippocampal CA1, the stratum pyramidale of the CA3, and the dentate gyrus decreased transiently in the early postischemic phase but then recovered. However, [3H]cyclic AMP binding in the strata pyramidale and radiatum of the CA1, the granular layer of the dentate gyrus, and the upper layer of the cortex decreased again 7 days after ischemia. In the CA4 subfield and the lower layer of the cortex, the binding showed no significant alterations after ischemia. Administration of pentobarbital prior to the induction of ischemia prevented the decrease in [3H]cyclic AMP binding in the CA1 subfield 6 h and 7 days after ischemia, and showed protective effects against neuronal death of the CA1 pyramidal cells 7 days after ischemia. These results indicate that marked alteration of intracellular signal transduction precedes neuronal damage in the hippocampal CA1 subfield. Furthermore, postischemic reduction of [3H]cyclic AMP binding in the histologically intact cerebral cortex, CA3, and dentate gyrus may be the reflection of cellular dysfunction after energy failure.     

Neuronal damage following non-lethal but repeated cerebral ischemia in the gerbil

Summary Brief, non-lethal transient forebrain ischemia in the gerbil can injure selectively vulnerable neurons when such ischemia is induced repeatedly. The influence of the number and interval of the ischemic insults on neuronal damage, as well as the time course of damage, following repeated 2-min forebrain ischemia were examined. A single 2-min forebrain ischemia were examined. A single 2-min ischemic insult caused no morphological neuronal damage. A moderate number of hippocampal CA1 neurons were destroyed following two ischemic insults with a 1-h interval, and destruction of almost all CA1 neurons resulted from three or five insults at 1-h intervals. Three and five insults also resulted in moderate to severe damage to the striatum and thalamus, depending on the number of episodes. Although three ischemic insults at 1-h intervals caused severe neuronal damage, this number of insults at 5-min and 4-h intervals caused destruction of relatively few neurons, and non neurons were destroyed at 12-h intervals. Following three ischemic insults at 1-h intervals, damage to the striatum, neocortex, hippocampal CA4 subfield and thalamus was observed at 6–24 h of survival, whereas damage to the hippocampal CA1 subfield appeared at 2–4 days. The results indicate that even a brief non-lethal ischemic insult can produce severe neuronal damage in selectively vulnerable regions when it is induced repeatedly at a certain interval. The severity of neuronal damage was dependent on the number and interval of ischemic episodes.     

The effects of maternal ethanol exposure on neurotransmission and second messenger systems: a quantitative autoradiographic study in the rat brain.

The effects of maternal ethanol exposure on neurotransmission and second messenger systems were examined in rats using histochemistry and in vitro autoradiography. Thirty % ethanol was administered to pregnant rats from gestational day 7 to the day of delivery. Quantitative autoradiography was used to map muscarinic cholinergic, dopamine D2, adenosine A1, and inositol 1,4,5-trisphosphate binding sites, as well as to localize adenylate cyclase and protein kinase C. We found no difference in the patterns of staining with acetylcholinesterase and Timm's stain between control and prenatally ethanol-exposed rats on postnatal day (PN) 30. In the ethanol-exposed rats, [3H]forskolin binding sites were increased during early development in the CA1 subfield of the hippocampus and the occipital cortex; [3H]phorbol ester binding sites were increased in the cortex, striatum, and hippocampus; hippocampal muscarinic cholinergic sites were increased on PN4 and 30; adenosine A1 binding was reduced on PN10 in most regions examined, but was increased in the CA1 subfield on PN30; dopamine D2 receptor levels were significantly reduced on PN30 in the striatum; and IP3 receptors were decreased in most regions studied, but particularly in the cerebellum. Thus, some of these changes were transient and others were long-lasting. Although histopathological abnormalities were minimal, the alterations of binding sites in the cerebellum (the coordination center) and in the hippocampus (related to memory and learning) that were detected may contribute to the behavioral and mental deterioration seen in the fetal alcohol syndrome.     

Neuronal damage and calcium accumulation following repeated brief cerebral ischemia in the gerbil

We investigated the distribution of neuronal damage following brief cerebral transient ischemia and repeated ischemia at 1-h intervals in the gerbil, using light microscopy and 45Ca autoradiography as a marker for detection of ischemic damage. The animals were allowed to survive for 7 days after ischemia induced by bilateral carotid artery occlusion. Following 2-min ischemia, neuronal damage determined by abnormal calcium accumulation was not observed in the forebrain regions. Following 3-min ischemia, however, abnormal calcium accumulation was recognized only in the hippocampal CA1 sector and part of the striatum. Two 2-min ischemic insults caused extensive abnormal calcium accumulation in the dorsolateral part of striatum, the hippocampal CA1 sector, the thalamus, the substantia nigra and the inferior colliculus. The ischemic insults were more severe than that of a single 3-min ischemia. However, three 1-min ischemic insults caused abnormal calcium accumulation only in the striatum. On the other hand, three 2-min ischemic insults caused severe abnormal calcium accumulation in the brain. The abnormal calcium accumulation was found in the dorsolateral part of striatum, the hippocampal CA1 sector, the thalamus, the medial geniculate body, the substantia nigra and the inferior colliculus. Gerbils subjected to three 3-min ischemic insults revealed most severe abnormal calcium accumulation. Marked calcium accumulation was seen not only in the above sites, but also spread in the neocortex, the septum and the hippocampal CA3 sector. Morphological study after transient or repeated ischemia indicated that the distribution and frequency of the neuronal damage was found in the sites corresponding to most of the regions of abnormal calcium accumulation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Mapping second messenger systems in the rat hippocampus after transient forebrain ischemia: in vitro [3H]forskolin and [3H]inositol 1,4,5-trisphosphate binding

Autoradiographic imaging demonstrated predominant and reciprocal localization of forskolin and inositol 1,4,5-trisphosphate (IP3) binding sites in synaptic areas in the hippocampus. We produced selective damage to the CA1 pyramidal cells in the rat hippocampus by means of transient forebrain ischemia and analyzed the alteration of the intracellular signal transduction using quantitative autoradiography of these second messenger systems. The dendritic fields (stratum oriens, radiatum and lacunosummoleculare) in the CA1 showed 20% decrease in [3H]IP3 binding activity 3 h after ischemia, when no morphological abnormalities were obvious. Thereafter, grain density in these layers decreased and half of the binding sites were lost 2 days after ischemia. By contrast, the stratum pyramidale of the CA1 showed no significant change until 2 days after recirculation. Seven days after ischemia, when CA1 pyramidal cells were depleted, all layers in the CA1 subfield lost 85% of [3H]IP3 binding sites. In the CA3 subfield, only a small and transient alteration in the [3H]IP3 binding was noticed during recirculation. Postischemic reduction of [3H]forskolin binding sites was obvious in the CA1 only 1 h after ischemia followed by loss of 50% of binding activity 7 days after recirculation. These results suggest that forskolin and IP3 binding sites are predominantly distributed on the pyramidal cells in the CA1 subfield and that marked alteration of intracellular signal transduction precedes the delayed CA1 pyramidal cell death.     

Repeated focal cerebral ischemia in gerbils is associated with development of infarction.

We induced repeated focal cerebral ischemia in gerbils. Single 5-min ischemia produced neuronal damage limited to the ipsilateral CA1 and CA4 hippocampus. Two 5-min ischemic insults spaced at a 1-h interval caused selective neuronal damage to the CA1, CA3 and CA4 hippocampus, striatum, neocortex, and thalamus. Three 5-min ischemic insults at 1-h intervals produced infarction. Thus, repeated focal ischemia produced cumulative brain damage by conversion of sublethal damage into selective neuronal damage and of the neuronal damage into infarction.     

Effects of naftidrofuryl oxalate, a 5HT2 antagonist, on neurotransmission and transduction systems in the gerbil hippocampus

We investigated the effects of age and naftidrofuryl oxalate (Naftidrofuryl), a 5-HT₂ antagonist, on neurotransmission and transduction systems in the gerbil hippocampus using quantitative autoradiography. [³H]Quinuclidinyl benzilate (QNB), [³H]cyclohexyl-adenosine (CHA), [³H]MK-801, and [³H]muscimol were used to label muscarinic acetylcholine, adenosine A1, N-methyl-d-aspartate (NMDA), and γ-aminobutyric acid-A (GABA_A) receptors, respectively. [³H]PN200-110 labeled L-type Ca²⁺ channels. [³H]Forskolin, [³H]cyclic adenosine monophosphate (cAMP), [³H]phorbol 12,13-dibutyrate (PDBu), and [³H]inositol 1,4,5-triphosphate (IP₃) were used to label adenylate cyclase, cAMP-dependent protein kinase, protein kinase C (PKC), and IP₃ receptors, respectively. Approximately 20% reductions in [³H]QNB, [³H]forskolin, and [³H]PDBu binding were observed in the hippocampus of 9-month-old gerbils in comparison with 5-week-old gerbils. Treatment with Naftidrofuryl (10 mg/kg, i.p., once a day for 7 days) ameliorated these reductions. No changes were found in [³H]CHA, [³H]MK-801, [³H]muscimol, [³H]PN200-110, [³H]cAMP, and [³H]IP₃ binding. The results suggest that Naftidrofuryl may have beneficial effects on the age-related alterations in signal transmission and transduction systems in the brain. Because the acetylcholine system, adenylate cyclase, and PKC are considered to be involved in learning and memory processes, the result may have clinical implications.     

Excitatory amino acid binding sites in the rat hippocampus after transient forebrain ischemia

The influence of transient forebrain ischemia on the temporal alteration of glutamate receptors in the hippocampal formation was analyzed by means of in vitro quantitative receptor autoradiography. We compared the binding of N-methyl-D-aspartate (NMDA) receptors using [3H]3-[+/-)2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid (CPP), noncompetitive NMDA antagonist binding sites using [3H]N-(1-(2-thienyl)-cyclohexyl)-3,4-piperidine (TCP), and kainate (KA) receptors. In the CA1 subfield of the hippocampus, the number of NMDA receptors and noncompetitive NMDA antagonist binding sites remained constant during the early stage of recirculation when the CA1 pyramidal cells remained histologically intact. A significant reduction of these receptor densities was observed 7 days following ischemia, when NMDA receptors and noncompetitive NMDA antagonist binding sites lost 64 and 29% of their binding sites in the stratum radiatum of the CA1, respectively. The KA receptor density in the CA1 subfield decreased by 44% 7 days after ischemia. Marked loss of the above-mentioned receptors in the CA1 after selective depletion of the CA1 pyramidal cells indicated that NMDA receptors, noncompetitive NMDA antagonist binding sites, and KA receptors in the CA1 are predominantly localized on the CA1 pyramidal cells. NMDA receptor density in the CA3 gradually decreased during the recirculation period. The stratum moleculare of the dentate gyrus, whose structure was histologically intact after ischemic insult, also showed a reduction of NMDA receptors 7 days following ischemia. [3H]KA receptor density in the stratum lucidum of the CA3 and in the hilus also decreased during recirculation. These     

Protective effect of cyclohexyladenosine on adenosine A1-receptors, guanine nucleotide and forskolin binding sites following transient brain ischemia: a quantitative autoradiographic study

The effects of an i.p. administration of cyclohexyladenosine (CHA) have been examined upon ischemic brain damage in gerbils. Ischemia was induced for 20 min by occlusion of both carotid arteries, and CHA was administered 5 min after recirculation at a dose of 2 mg/kg. Animals were sacrificed either 1, 3 or 6 days after ischemia and their brains were used for examination of cell morphology and quantitative autoradiography. In animals subject to ischemia, the deterioration of the laminar organization of the hippocampus was associated with a significant decrease in adenosine A1-receptors (labeled with [3H]CHA), G-protein (labeled with [3H]forskolin). The treatment with CHA considerably improved the morphological preservation of cells in the CA1 region of the hippocampus and prevented the reduction in the specific binding of all radioligands. Adenosine, its analogues and other substances modulating adenosine receptors may thus provide new therapeutic approaches to the treatment of ischemia-induced brain injury.     

Emphasized selective vulnerability after repeated nonlethal cerebral ischemic insults in rats.

BACKGROUND AND PURPOSE: We examined the density and distribution of brain damage after repeated periods of nonlethal ischemic insult in rats in comparison with damage after single lethal periods of ischemic insult. METHODS: Transient cerebral ischemia was induced by four-vessel occlusion for 3, 10, 20, and 30 minutes, and 3-minute periods of ischemia were repeated two, three, or five times at 1-hour intervals, followed by 7 days of survival. RESULTS: Three minutes of ischemia produced no brain damage, but 10-30 minutes of ischemia produced neuronal damage, depending on the length of ischemia, to the selectively vulnerable forebrain regions such as hippocampal CA1 and CA4 subfields, neocortex, striatum, and ventral thalamus, as well as to the brain stem structures (medial geniculate body, substantia nigra, and inferior colliculus) and cerebellar Purkinje cells. Two 3-minute periods of ischemic insult produced neuronal damage to the hippocampal CA1 subfield. Three and five 3-minute insults produced neuronal damage extensively to the selectively vulnerable forebrain areas. An intense cumulative effect of damage was observed in the ventral thalamus, whereas the substantia nigra and the inferior colliculus were resistant to repeated ischemic insults. CONCLUSIONS: Our data indicate that the density and distribution of neuronal damage after repeated ischemic insults are altered as compared with after single ischemia.     

Effects of hyperthermia on the effectiveness of MK-801 treatment in the gerbil hippocampus following transient forebrain ischemia

The effects of dizocilipine maleate (MK-801), a noncompetitive N-methyl-D-aspartate (NMDA) receptor/channel antagonist, were tested on the dysfunction of neurotransmitter and signal transduction systems and morphological damage 7 days after transient forebrain ischemia in gerbils. Neurotransmitter system (adenosine A1, muscarinic cholinergic receptor) and signal transduction system (inositol 1,4,5-trisphosphate receptor: IP3, protein kinase C: PKC, L-type calcium channels) binding sites were mapped by in vitro quantitative receptor autoradiography. All ligands used in the present study decreased significantly in the CA1 subfield 7 days after ischemia. In normothermic animals, pretreatment with MK-801 failed to protect against decreased receptor binding in the hippocampus 7 days after ischemia. Moreover, in a morphological study, pre- and posttreatment of MK-801 failed to show protective effects against ischemic neuronal damage. On the other hand, pretreatment of MK-801, without maintaining body temperature, prevented the neuronal death of CA1 subfield 7 days after ischemia. These results weaken the hypothesis that NMDA receptor/channel may play a pivotal role in the pathogenesis of neuronal damage after transient forebrain ischemia.