Intracerebroventricular Injection Of Colchicine Research Articles

Contemporary comprehensive approaches to assessing the effectiveness of experimental model of neurodegenerative disorders with cognitive status changes

Aim of the study was to characterize the locomotor and cognitive aspects of the behavior of experimental rats under intracerebroventricular colchicine administration in the open field test and the 8-arm radial maze, as well as identify a set of behavioral features of experimental animals that emerged during the study.
 Materials and methods. The study was conducted in two stages on 20 male Wistar rats aged 10–11 months. The first stage involved assessment of initial locomotor activity and cognitive functions in all intact rats. At the next stage, the rats were divided into two experimental groups (n = 10): the first group with intracerebroventricular injection of physiological NaCl solution, and the second group with intracerebroventricular injection of colchicine. In 14 days after the surgery, repeated recording of locomotive and cognitive activity indicators was performed.
 Results. The locomotor activity characteristics did not statistically differ between the animals before the surgical procedures and the rats of the first group. However, in the second group, the activity indicators were significantly higher than in the respective pre-surgery rats. In the intergroup comparison of the rats that entered the second stage of the experiment, it was found that among all the investigated parameters, only the indicator of high activity duration was significantly higher in the second group compared to the first group. At the same time, the animals in the second group showed significant cognitive impairments compared to the first group, as indicated by significantly lower memory index values, the number of correct entries into the maze arms, and a significantly longer time to make the first correct entry into the maze arm.
 Conclusions. Intracerebroventricular administration of colchicine to experimental rats is accompanied by increased locomotor activity and impairment of cognitive functions. The administration of a physiological solution is not accompanied by a statistically significant increase in locomotor activity, but it demonstrates a clear tendency to increase, which may indicate a certain influence of the procedure itself. The applied pharmacological model of neurodegeneration with subsequent comprehensive assessment of animal behavior in an open field and an 8-arm radial maze is legitimate and can be used to study the early development of neuroinflammation, neuroapoptosis, and synaptogenesis disorders in the experiment.

Effect of sacubitril/valsartan on cognitive impairment in colchicine-induced Alzheimer's model in rats.

Alzheimer's disease (AD) is a complex neurodegenerative disease. There is epidemiological evidence that heart failure (HF) patients are at higher risk of developing AD, and the impact of sacubitril/valsartan, the first angiotensin receptor-neprilysin inhibitor (ARNI) approved for HF, on cognitive functions is still controversial. To investigate the effect of sacubitril/valsartan on cognitive functions in colchicine-induced AD rat model. Forty adult male Wistar rats were equally allocated into four groups (each of 10 rats): Group I: normal control, Group II: intracerebroventricular injection of colchicine (15 μg/5μl/bilaterally), Group III: colchicine (15 μg/5μl/bilaterally, icv) + oral sacubitril/valsartan (100 mg/kg/day) for 25 days, and Group IV: colchicine (15 μg/5μl/bilaterally, icv) + oral valsartan (50 mg/kg/day) for 25 days. Behavioral assessment was done using Morris water maze and passive avoidance tasks. Biochemically, β-amyloid (1-40 and 1-42) peptides, oxidative stress (malondialdehyde and superoxide dismutase) and inflammatory (tumor necrosis factor-alpha) parameters were measured in hippocampus and prefrontal cortex. Sacubitril/valsartan exaggerated colchicine-induced cognitive impairment in both Morris water maze and passive avoidance tasks and was associated with significant increase in β-amyloid accumulation, oxidative stress, and inflammation versus valsartan. Sacubitril/valsartan caused deleterious effect on cognitive impairment and biochemical alterations in colchicine-induced AD rat model. Hence, special caution should be taken following long-term intake of ARNI on cognitive functions.

Protective Role of Caffeic Acid in Cognitive Dysfunction and Oxidative Stress Induced by Colchicine in Rats

Purpose: Alzheimer’s is a disease affecting mostly the Older population leading to the deterioration of cognitive capabilities. The protective effect of Caffeic acid in Colchicineinduced dementia was evaluated in the current study. Materials and Methods: Colchicinewas administered intracerebroventricularly (ICV) to the lateral ventricle of the brain (at the coordinates 0.8 mm posterior to bregma, 1.8 mm lateral to the sagittal suture, 3.6 mm below the cortical surface) using robotic stereotaxic apparatus that results in Alzheimer’s type sporadic dementia. Caffeic acid at the dose of 50 mg/kg p.o, was administered daily for 25 days starting four days before the colchicine injection and evaluated for its neuroprotective activity. The spatial memory of animals was evaluated using Morris water maze followed by biochemical estimations of acetylcholinesterase and antioxidant markers in the hippocampal and frontal cortex region of the brain. Results: Intracerebroventricular injection of colchicine in rat brain resulted in decreased cognitive abilities as evident in escape latency and average speed of the retention trial. Significant changes in the escape latency were noted in Caffeic acid-treated group. The level of acetylcholinesterase and antioxidant markers like glutathione, catalase, lipid peroxidation, superoxide dismutase were significantly changed in the hippocampal region of the rats but not in the frontal cortex region in the caffeic acid treatment groups. Conclusion: The current study provides evidence for the neuroprotective and antioxidative potential of caffeic acid in intracerebroventricularly injected Colchicine-induced sporadic model of AD.

Co‐expression of Orexin‐B and Corticotropin‐Releasing Factor in Hypothalamic Neurons of the Ovariectomized Female on a High Fat Diet

The hypothalamic paraventricuar nucleus (PVH) is known to play a pivotal role in energy balance and in autonomic and neuroendocrine control. The neuropeptides orexin‐A (OX‐A) and orexin‐B (OX‐B) have been implicated in these homeostatic mechanisms. This study was done to investigate whether 17β‐estradiol (E) treatment coupled with ingestion of a high fat diet (HFD; 45 kcal fat) altered OX immunoreactivity in PVH of ovariectomized (OVX) female Wistar rats treated (OVX+E) or not‐treated with E (OVX only). Animals were placed on a normal diet (ND) or HFD for a period of 7 or 20 weeks. Twenty‐four hours before sacrifice all animals were given an intracerebroventricular injection of colchicine and the forebrains were processed for OX‐A‐and OX‐B‐ immunoreactivity. In all cases, relative OX‐B immunoreactive fiber density was greater throughout PVH compared to OX‐A in both female groups regardless of the diet. OX‐A‐immunoreactivity was also found to be greater in females fed a HFD compared to a ND. Furthermore, a distinct cluster of OX‐B immunoreactive neurons was found within the medial parvocellular PVH in OVX only animals, and the number of these neurons was found to higher in those fed a HFD for the longer period. Pre‐pro‐orexin protein expression was up‐regulated in PVH in OVX only females on the HFD compared to ND and compare to OVX+E on a HFD or ND. As the location of these OX‐B neurons in the medial parvocellular PVH overlapped the area known to contain corticotropin‐releasing factor (CRF) neurons, a double immunofluorescence studied was done to determine whether the OX‐B neurons co‐expressed CRF. All medial parvocellular PVH neurons that expressed OX‐B co‐expressed CRF, while not all neurons expressing CRF co‐expressed OX‐B. As OX has been shown to function as a neuromodulatory hormone that stimulates the adrenal corticotrophin hormone‐mediated response to stress, these findings suggest that in females without E, co‐release of OX‐B and CRF in response to a HFD stressor may affect anterior pituitary function.Support or Funding InformationSupported in part by the Heart and Stroke Foundation.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

A comparison of neurodegeneration linked with neuroinflammation in different brain areas of rats after intracerebroventricular colchicine injection

Colchicine induces neurodegeneration, but the extent of neurodegeneration in different areas of the brain in relation to neuroinflammation remains unclear. Such information may be useful to allow for the development of a model to compare colchicine-induced neurodegeneration with other neurodegenerative diseases such as Alzheimer’s Disease (AD). The present study was designed to investigate the extent of neurodegeneration along with neuroinflammation in different areas of the brain, e.g. frontal cortex, parietal cortex, occipital cortex, corpus striatum, amygdala and hippocampus, in rats along with memory impairment 21 days after a single intracerebroventricular (icv) injection of colchicine. Memory parameters were measured before and after icv colchicine injection in all test groups of rats (control, sham-operated, colchicine-injected [ICIR] rats). On Day 21 post-injection, rats from all groups were anesthesized and tissues from the various brain areas were collected for assessment of biomarkers of neuroinflammation (i.e. levels of ROS, nitrite and proinflammatory cytokines TNFα and IL-1β) and neurodegeneration (assessed histologically). The single injection of colchicine resulted in impaired memory and neurodegeneration (significant presence of plaques, Nissl granule chromatolysis) in various brain areas (frontal cortex, amygdala, parietal cortex, corpus striatum), with maximum severity in the hippocampus. While IL-1β, TNFα, ROS and nitrite levels were altered in different brain areas in the ICIR rats, these parameters had their greatest change in the hippocampus. This study showed that icv injection of colchicine caused strong neurodegeneration and neuroinflammation in the hippocampus of rats and the increases in neurodegeneration were corroborated with those of neuroinflammation at the site. The present study also showed that the extent of neurodegeneration and neuroinflammation in different brain areas of the colchicine-injected rats were AD-like and supported the fact that such rats might have the ability to serve as a sporadic model of AD.

Effects of Naproxen on Immune Responses in a Colchicine-Induced Rat Model of Alzheimer's Disease

Background: The components of the immune system have been indicated to be linked with the neurotoxicity in Alzheimer's disease (AD). The participation of the immune system in the neurodegeneration in a rat model of colchicine-induced AD has not been explored. Methods: In the present study, hippocampal neurodegeneration along with reactive oxygen species (ROS), nitrite and TNF-α in the hippocampus and some systemic immune responses were measured after 15 and 21 days of intracerebroventricular colchicine injection in rats and again after oral administration of different doses of the anti-inflammatory drug naproxen in AD rats. Results: Chromatolysis and amyloid plaques were found along with higher ROS, nitrite and TNF-α levels in the hippocampus of colchicine-induced AD rats, and these changes were prevented by naproxen in a dose-dependent manner. Alterations in immunological parameters [increased phagocytic activity of white blood cells and splenic polymorphonuclear cells (PMN), increased cytotoxicity and decreased leucocyte adhesive inhibition index (LAI) of splenic mononuclear cells (MNC)] were also observed in colchicine-injected rats, which showed a dose-dependent recovery after oral administration of naproxen in AD rats. The number of plaques, chromatolysis of Nissl granules, TNF-α, nitrite and ROS levels in the hippocampus, phagocytic activity of splenic PMN and LAI of splenic MNC in AD rats showed greater changes in the 21- than in the 15-day study, and the recovery of these parameters after administration of naproxen differed between the two study durations. Conclusion: The present study shows that colchicine-induced neurodegeneration is time dependent and mediated by cyclooxygenase-induced neuroinflammation, which is reflected in the systemic immunological responses.

High fat diet impact on Fos expression in ovariectomized female C57BL/6 mice: effect of colchicine and response of different neuronal phenotypes

Activity of neuropeptide Y (NPY), tyrosine hydroxylase (TH), corticoliberine (CRH), and oxytocin (OXY) producing cells was investigated in the ovariectomized (OVX) female C57BL/6 mice kept on the high fat diet for 16 weeks and their response to colchicine stress in selected brain areas, including the hypothalamic paraventricular (PVN), dorsomedial (DMN) and arcuate (ARC) nuclei, A1/C1 (in the ventrolateral medulla), and A2/C2 (in the nucleus of the solitarii tract, NTS) catecholaminergic cell groups. The OVX female C57BL/6 mice kept on high fat diet were sacrificed by transcardial perfusion with fixative 48 h after intracerebroventricular injection of colchicine (18 µg mice). Dual Fos/neuropeptide immunohistochemistry was employed to investigate Fos/neuropeptide colocalizations. In the OVX saline-treated mice (sham control) with standard diet (St diet), no immunopositive CRH and NPY neurons were identified in the PVN and weak Fos immunostainig was visible in TH neurons in the DMN and ARC nuclei. Colchicine treatment in the OVX mice with St diet increased the number of CRH and OXY immunopositive neurons in the PVN as well as the number of NPY and TH neurons in DMN and ARC nuclei and NPY neurons in the middle NTS (mNTS) and A1/C1 cell group. Prolonged HF diet in OVX sham control mice moderately increased the number of Fos/TH neurons in the mNTS and commissural NTS (cNTS) in comparison with St diet mice. However, prolonged HF diet in OVX colchicines-treated mice reduced the number of Fos/NPY neurons in the anterior NTS (aNTS) and A1/C1 cell group in comparison with colchicines-treated animals with St diet as well as Fos-TH neurons in the mNTS and cNTS in comparison with saline-treated animals with HF diet. The data of this pilot study indicate that prolonged high fat diet might: 1) represent itself a light/moderate stimulus for activation of TH neurons in the NTS and A1/C1 cell group as well as NPY neurons in the A1/C1 cell group and 2) interfere with colchicines-induced and time-delayed Fos activation in the NPY and TH neurons in both the above mentioned brain nuclei.

Morphofunctional interactions between galanin and GnRH-containing neurones in the diencephalon of the ewe. The effect of oestradiol

In rodents, the neuropeptide galanin (Gal) is involved in controlling the release of gonadotrophin-releasing hormone (GnRH). In the female, this peptide is colocalized in a subpopulation of GnRH neurones and its expression is stimulated by oestradiol. In the ewe, the morphofunctional relationship between these two neuronal peptides is poorly understood. The morphological interaction between Gal and GnRH was studied in ewes treated with oestradiol or with colchicine and in control animals. Five ewes were treated for 6h with oestradiol implants, a treatment known to induce a preovulatory surge of GnRH, and compared with five control animals. In addition, four animals received an intracerebroventricular injection of colchicine known to increase the intracellular level of galanin immunoreactivity. The morphological relationship between the two peptides was investigated by immunofluorescence using specific antibodies on the same sections, and the results were analysed using confocal microscopy. In colchicine-treated ewes, numerous Gal-immunoreactive neurones were found in the preoptic area in the vicinity of GnRH-immunoreactive neurones, but the two peptides were never observed in the same neurone. In all animals, Gal-ir fibres were observed to be in apposition to GnRH-containing perikarya in the preoptic area and these appositions were more numerous in oestradiol-treated ewes than in control animals. In contrast with rodents, galanin was not colocalized with GnRH in the neurones of the preoptic area of ewes, but this peptide could control GnRH neuronal secretion through axosomatic interactions. However, the presence of synaptic contacts between galanin terminals and GnRH perikarya needs to be confirmed by electron microscopy. As in rodents and primates, galanin could mediate the positive feedback of oestradiol on GnRH neurones during the preovulatory surge in ewes.

Neuroprotective Effects of Resveratrol against Intracerebroventricular Colchicine-Induced Cognitive Impairment and Oxidative Stress in Rats

Alzheimer’s disease is a complex and multifactorial neurodegenerative disease. Central administration of colchicine, a microtubule-disrupting agent, causes loss of cholinergic neurons and cognitive dysfunction that is associated with excessive free radical generation. The present study was aimed at evaluating the effects of trans-resveratrol in the prevention of colchicine-induced cognitive impairment and oxidative stress in rats. Intracerebroventricular administration of colchicine (15 µg/5 µl) induced impaired cognitive functions in both the Morris water maze task and the elevated plus-maze task. Chronic treatment with resveratrol (10 and 20 mg/kg, p.o.) for a period of 25 days, beginning 4 days prior to colchicine injection, significantly improved the colchicine-induced cognitive impairment. Intracerebroventricular colchicine injection resulted in free radical generation characterized by alterations in oxidative stress markers with a significant increase in malondialdehyde (MDA) and nitrite levels and depletion of reduced glutathione (GSH) activity in the rat brains. It also showed a significant decrease in acetylcholinesterase activity. Besides improving cognitive dysfunction, chronic administration of resveratrol significantly reduced the elevated MDA and nitrite levels and restored the depleted GSH and acetylcholinesterase activity. Results of the present study indicated that trans-resveratrol has a neuroprotective role against colchicine-induced cognitive impairment and associated oxidative stress.

Differential effects of cyclooxygenase inhibitors on intracerebroventricular colchicine-induced dysfunction and oxidative stress in rats

Alzheimer's disease is a progressive neurological and psychiatric disorder. Oxidative stress and neuroinflammation have been implicated in pathophysiology of Alzheimer's disease. Inflammatory cells, such as astrocytes and microglia, are activated in areas of the brain affected by amyloid plaques and inflammatory mediators including cytokines, chemokines, prostaglandins, oxygen free radicals and reactive nitrogen species may have a crucial role in Alzheimer's disease pathogenesis. Central administration of colchicine, a microtubule-disrupting agent, causes loss of cholinergic neurons and cognitive dysfunction that is associated with excessive free radical generation. The present study was aimed to evaluate the effects of cyclooxygenase inhibitors against colchicine-induced cognitive dysfunction and oxidative stress in rats. Following intracerebroventricular (i.c.v.) administration of colchicine (15 μg/5 μl), rats exhibited poor retention of memory in Morris water maze and elevated plus maze task paradigms and oxidative stress in rats. Chronic treatment with naproxen ( per se; 20 and 40 mg/kg, p.o.) or valdecoxib ( per se; 5 and 10 mg/kg, p.o.) daily respectively for a period of 25 days beginning 4 days prior to colchicine injection significantly improved colchicine-induced cognitive impairment. Intracerebroventricular colchicine injection resulted in free radical generation characterized by alterations in oxidative stress markers with a significant increase in malondialdehyde and nitrite levels and depletion of reduced glutathione levels in the brains of rats. It also caused a decrease in acetylcholinesterase activity. Besides, improving cognitive dysfunction, chronic administration of cyclooxygenase inhibitors (naproxen and valdecoxib) significantly reduced elevated malondialdehyde, nitrite levels and restored reduced glutathione levels and acetylcholinesterase activity. The results of the present study indicated that naproxen ( per se; 20 and 40 mg/kg, p.o.) or valdecoxib ( per se; 5 and 10 mg/kg, p.o.) treatment has a neuroprotective role against colchicine-induced cognitive impairment and associated oxidative stress. The present findings further support the potential use of cyclooxygenase inhibitors in treatment of neurodegenerative diseases such as Alzheimer's disease.

Interleukin-6 and Nitric Oxide Synthase Expression in the Vasopressin and Corticotrophin-releasing Factor Systems of the Rat Hypothalamus

Nitric oxide synthase (NOS) and interleukin-6 (IL-6) are constitutively expressed in hypothalamic cells. However, phenotypic and functional aspects of these cells remain unknown. We have studied the expression pattern of these two molecules in hypothalamic cells expressing corticotropin-releasing factor (CRF) and arginin-vasopressin (AVP), two major regulatory peptides in the hypothalamus-pituitary system, using immunofluorescence, intracerebroventricular injection of colchicine, and the study in parallel of the labeling pattern of axons in the median eminence. Within AVP cells, we distinguished two different populations: large, intensely stained AVP cells coexpressing IL-6; and large, intensely stained AVP cells coexpressing IL-6 and NOS. Within the CRF cells, we distinguished three different populations: large, intensely stained CRF cells immunonegative for AVP, NOS, and IL-6; large cells weakly stained for CRF and AVP, immunopositive for NOS and immunonegative for IL-6; and small cells intensely stained for CRF and AVP and immunonegative for IL-6 and NOS. In addition, we also found AVP cells containing IL-6 in the suprachiasmatic nucleus. These results suggest that neuronal NOS and IL-6 may be involved in different modulatory processes in hypophysiotropic and non-hypophysiotropic cells.

Differential effects of colchicine on the induction of nitric oxide synthase in neurons containing progesterone receptors of the guinea pig hypothalamus

Using nicotinamide-adenine-dinucleotide-phosphate-diaphorase (NADPHd) histochemistry, we analyzed the effects of an intracerebroventricular injection of colchicine on the activity of neuronal nitric oxide synthase in the hypothalamic nuclei of intact and ovariectomized estradiol-primed guinea pigs. We also examined the effects of colchicine on the immunocytochemical colocalization of nitric oxide synthase with the progesterone receptor in the ventrolateral nucleus, a key region in the control of sexual behavior. Treatment with colchicine resulted in a significant increase in the number of NADPHd-positive neurons in the ventrolateral nucleus in intact as well as in ovariectomized estradiol-primed animals, whereas in the other hypothalamic regions analyzed (preoptic area, paraventricular nucleus and posterior arcuate nucleus), the enzymatic activity remained unchanged. Quantitative analysis showed a significantly greater number of NADPHd-positive cells in the medial and the posterior aspects of the ventrolateral nucleus of colchicine-treated guinea pigs compared to the control group. In the caudal subdivision of this nucleus, colchicine induced nitric oxide synthase in the target cells for progesterone. These results suggest that neuronal nitric oxide synthase activity in the hypothalamus is affected by colchicine in a region-specific manner and especially in the ventrolateral nucleus, which is involved in progesterone-facilitated lordosis.

Colchicine Differentially Induces the Expressions of Nitric Oxide Synthases in Central and Peripheral Catecholaminergic Neurons

This study was aimed at elucidating differences in nerve injury induced expression of nitric oxide synthases (NOS) between the peripheral and central catecholaminergic neurons. Colchicine was used to disrupt chemically the neuronal cytoskeletal integrity. A marked increase in the expression of neuronal NOS-IR and NADPH-diaphorase activity, a marker of neuronal NOS (nNOS), was seen in distinct populations of postganglionic sympathetic neurons of the superior cervical ganglion after intraganglionic colchicine injection. Similarly, immunoreactivity for the inducible form of NOS (iNOS) was induced in some sympathetic neuron somata. However, this immunoreactivity did not coincide with nNOS-IR. In contrast to the sympathetic neurons, hypothalamic arcuate and periventricular dopaminergic neurons did not show NOS-IR or NADPH-DA either in intact animals or in animals treated with an intracerebroventricular injection of colchicine. Immunoreactivity for the inducible form of NOS revealed no neuronal staining in the hypothalamic neurons in either group, while a large number of glia-resembling cells around the third ventricle showed slight expression of iNOS-IR. The present results show that expression of both neuronal and inducible forms of NOS may be induced by colchicine in some catecholaminergic neurons. It is suggested that these inductions are specific to certain catecholaminergic neuronal systems, like the sympathetic neurons, rather than a general property of catecholaminergic neurons.

Colchicine affects cortical and amygdalar neurochemical changes differentially after middle cerebral artery occlusion in rats

Recently, we have shown increases in the immunoreactivity for neuropeptide Y and tyrosine hydroxylase in the insular cortex surrounding the focal infarction after middle cerebral artery occlusion. In addition, the immunoreactivity for neuropeptide Y, leucine-enkephalin, dynorphin, and neurotensin increased ipsilaterally in the amygdala. Increases in immunoreactivity were observed in nerve terminals and fibers; changes in the neuropeptides were maximal 3 days after stroke. Local excitotoxic injury of the insular cortex also elicited similar neuropeptide changes unilaterally in the same regions. In this study, immunohistochemistry was used following intracerebroventricular injection of colchicine and stroke to determine whether blockade of axonal transport would prevent these neurochemical changes. These experiments would also locate the putative cellular origins of the neurochemicals involved. Control rats received either colchicine injection or middle cerebral artery occlusion alone. Injection of colchicine enhanced the periinfarct increase in neuropeptide Y but did not alter the increase in tyrosine hydroxylase. The neuropeptide Y increase was observed in local cortical neurons. Colchicine prevented the increases in immunoreactivity for the neuropeptides in the amygdala on the side of stroke, although there were small perikarya that showed immunoreactivity for these neuropeptides within the amygdala on both sides. We conclude that local cortical neurons are responsible for the increase in neuropeptide Y in the periinfarct region, that the cortical increase in tyrosine hydroxylase is not dependent on fast axonal transport, and that axonal transport of signals from the insular cortex to the amygdala is critical in mediating the amygdalar neuropeptide changes seen after stroke. J. Comp. Neurol. 387:27–41, 1997. © 1997 Wiley-Liss, Inc.

Colchicine affects cortical and amygdalar neurochemical changes differentially after middle cerebral artery occlusion in rats

Recently, we have shown increases in the immunoreactivity for neuropeptide Y and tyrosine hydroxylase in the insular cortex surrounding the focal infarction after middle cerebral artery occlusion. In addition, the immunoreactivity for neuropeptide Y, leucine-enkephalin, dynorphin, and neurotensin increased ipsilaterally in the amygdala. Increases in immunoreactivity were observed in nerve terminals and fibers; changes in the neuropeptides were maximal 3 days after stroke. Local excitotoxic injury of the insular cortex also elicited similar neuropeptide changes unilaterally in the same regions. In this study, immunohistochemistry was used following intracerebroventricular injection of colchicine and stroke to determine whether blockade of axonal transport would prevent these neurochemical changes. These experiments would also locate the putative cellular origins of the neurochemicals involved. Control rats received either colchicine injection or middle cerebral artery occlusion alone. Injection of colchicine enhanced the periinfarct increase in neuropeptide Y but did not alter the increase in tyrosine hydroxylase. The neuropeptide Y increase was observed in local cortical neurons. Colchicine prevented the increases in immunoreactivity for the neuropeptides in the amygdala on the side of stroke, although there were small perikarya that showed immunoreactivity for these neuropeptides within the amygdala on both sides. We conclude that local cortical neurons are responsible for the increase in neuropeptide Y in the periinfarct region, that the cortical increase in tyrosine hydroxylase is not dependent on fast axonal transport, and that axonal transport of signals from the insular cortex to the amygdala is critical in mediating the amygdalar neuropeptide changes seen after stroke.

Expression of Nitric Oxide Synthase in Hypothalamic Nuclei Following Axonal Injury or Colchicine Treatment

Nitric oxide (NO) has recently gained much attention due to its apparently double-edged role in neuronal injury. This study was aimed at elucidating neuronal nitric oxide synthase (nNOS) expression in the brain after two types of injury, namely axonal transection and colchicine treatment. The neurosecretory hypothalamo-pituitary pathway served as a model for the reaction of central neurons to these two types of injury. Axonal transection, i.e., pituitary stalk section, resulted in a qualitative increase in NOS content in the supraoptic and paraventricular nuclei. In these nuclei, there was also an increase in the number of NOS-expressing neurons after the operation. Surprisingly, in the periventricular nucleus, a strong decrease in the number of NOS-positive magnocellular neurons was observed in the anterior part of the nucleus. Intracerebroventricular injection of colchicine resulted in an increase in the cell count in the paraventricular nucleus, while the other nuclei remained unchanged. Our results suggest that axonal injury results in an increase in nNOS expression in the major neurosecretory nuclei, while the periventricular nucleus shows the opposite reaction. Colchicine treatment has an effect similar to that of axotomy in the major neurosecretory nuclei, suggesting that an increase in NOS expression may be induced by interrupted axonal transport. In the periventricular nucleus, the decrease in the number of NOS-containing neurons suggests differences among hypothalamic NOS-containing neuron groups in response to neuronal injury.

Expression of FGF-2 and FGF receptor type 1 in the adult rat brainstem: effect of colchicine.

In the adult rat brainstem, neuronal subpopulations of several motor and sensory nuclei display basic fibroblast growth factor (bFGF or FGF-2) immunoreactivity (IR; Grothe et al. J. Comp. Neurol. 305:328-336). In the present study we demonstrate that FGF-2-IR correlates with staining for the high-affinity FGF-receptor 1. Intracerebroventricular injection of colchicine leads to the disappearance or substantial reduction of FGF-2-IR in the hypoglossal, facial, trigeminal motor, trochlear, and mesencephalic trigeminal nuclei. In contrast, FGF-2-IR appears in many perikarya of the red nucleus and the medial nucleus of the trapezoid body, whereas in control rats both nuclei showed immunostained fibers and almost no immunoreactive cell bodies. This dramatic change of FGF-2-IR could be explained by the ability of colchicine to block fast axonal transport. Cranial nuclei may internalize FGF-2 at the periphery via high-affinity receptors and retrogradely transport the molecule to their perikarya. The red nucleus and the medial nucleus of the trapezoid body may synthesize FGF-2 and provide the growth factor to afferent or efferent neurons. The presence of FGF-2 mRNA in brainstem extract and the absence of the FGF-2 transcript in extracts of the hypoglossal nucleus corroborate this suggestion. The effect of colchicine on FGF-2-IR in the brainstem nuclei suggests that FGF-2 could be specifically retrogradely transported in other cranial nuclei, in addition to the hypoglossal system. Together with the ability of FGF-2 to stimulate neuronal survival, this result strongly supports the hypothesis that FGF-2 is acting as a neurotrophic factor on specific central neuron populations.

Relationship between neuropeptide Y and luteinizing-hormone-releasing hormone immunoreactivities in the hypothalamus and preoptic region.

The purpose of this study was to examine the anatomical relationships of perikarya and fibers containing neuropeptide Y (NPY) and luteinizing-hormone-releasing hormone (LHRH) in the hypothalamus and preoptic region of female rats. In view of our previous report of stimulatory effects of estrogen on LHRH and NPY levels in the median eminence, animals were bilaterally ovariectomized and subsequently implanted subcutaneously with capsules containing estradiol benzoate in oil or vehicle. Following intracerebroventricular injection of colchicine, rats were perfused with fixative and their brains sectioned and processed for immunohistochemical visualization of NPY and LHRH in the same section and in consecutive sections. Estrogen treatment had no discernible effect on the distribution or relationship of these peptides. NPY-immunoreactive fibers were intimately associated with LHRH-labeled primary dendrites and perikarya in the medial preoptic region and horizontal limb of the diagonal band of Broca. Fibers containing NPY or LHRH overlapped extensively in the lateral palisade region of the median eminence and also in the subependymal and internal zones. The external zone of the median eminence displayed relatively less overlap of these peptide systems. LHRH-immunoreactive axons coursed among NPY-labeled perikarya in the arcuate nucleus and appeared to contact these cells. These results suggest that NPY-containing axons may influence LHRH-positive neurons at the cell body and also at the site of axon termination in the median eminence. LHRH-containing axons appear to contact NPY-immunoreactive perikarya in the arcuate nucleus and may interact with terminals in the median eminence. This arrangement may provide a mechanism for communication between NPY and LHRH neurons and for the neuroendocrine coordination of hypothalamic NPY and LHRH secretion before ovulation.

Immunoreactivities for antisera to three putative neuropeptides of the rat melanin-concentrating hormone precursor are coexpressed in neurons of the rat lateral dorsal hypothalamus

Antisera (AS) raised against rat melanin-concentrating hormone (rMCH) and against two additional peptides sequences derived from the rat MCH precursor (neuropeptide glutamic acid-isoleucineamide (NEI), and neuropeptide glycine-glutamic acid (NGE)) exclusively stained the hypothalamic neurons previously described using AS to salmon MCH, human somatocrinin 1–37 (GRF37) and α-MSH. Liquid phase and dot-blot controls for specificity indicated that rMCH-, NEI- and NGE-AS bound epitopes recognized by sMCH-, α-MSH- and GRF-37-AS, respectively. The distinct intracellular patterns of immunoreactivity obtained in control animals with rMCH-, NGE- and NEI-AS, as well as the changes observed after intracerebroventricular injection of colchicine matched previous findings using sMCH-, GRF37- and α-MSH-AS.

Distribution of a renin-releasing factor in the central nervous system of the rat.

We have previously shown that the serotonergic regulation of renin secretion from the kidneys is mediated by a renin-releasing factor (RRF) that is present in both plasma and hypothalamus. The present studies were designed to determine the distribution of RRF in the brain and peripheral tissues and to test whether RRF release could be stimulated in vitro from hypothalamo-hypophyseal explants. RRF levels were determined in vitro by measuring renin release from kidney cortical slices. Addition of hypothalamic extract to rat kidney slices produced a dose-dependent increase in renin release. RRF was measurable in most brain areas with the highest renin-releasing activity in the hypothalamus, cerebral cortex, medulla oblongata and cerebellum. To determine which brain regions contain RRF cell bodies, rats received an intracerebroventricular injection of colchicine to inhibit axonal transport and concentrate RRF in the perikarya. After colchicine treatment, RRF activity in the cerebral cortex, medulla oblongata and cerebellum decreased. In contrast, the hypothalamus had increased RRF activity suggesting that RRF cell bodies are localized in the hypothalamus. Superfusion of hypothalamo-hypophyseal explants with a high potassium Krebs-Ringer solution stimulated RRF release, suggesting that depolarization of hypothalamic neurons can stimulate RRF secretion. Nephrectomy produced a significant increase in RRF concentration in the hypothalamus, suggesting that RRF neurons respond to decreased renin activity or other kidney-related substances in the circulation. The determination of RRF in peripheral tissue revealed minimal renin-releasing activity in the liver, spleen and skeletal muscle extracts. High performance chromatography of hypothalamic extract on a GPC-100 column revealed RRF activity in fractions that were estimated to have a molecular weight of 5,000. These studies suggest that RRF-containing cell bodies in the hypothalamus respond to depolarization by releasing RRF into the circulation. In addition, the hypothalamic content of RRF is regulated by the kidney. Altogether, these data suggest that RRF neurons are part of a neuroendocrine system that regulates renin secretion from the kidneys.