Temporomandibular inflammation regulates the matrix metalloproteinases MMP-2 and MMP-9 in limbic structures.

Temporomandibular disorder (TMD) is characterized by acute or chronic orofacial pain, which can be associated with inflammatory processes in the temporomandibular joint (TMJ) and emotional disorders. Peripheral and central sensitization in painful orofacial processes is common, and it can be triggered by peripheral inflammatory challenge with consequent neuroinflammation phenomena. Such neuroinflammation comes from inflammatory products from supportive cells, blood-brain barrier, and extracellular matrix. Here, we evaluated the possible recruitment of limbic structures for modified matrix metalloproteinases (MMPs) expression and activity during temporomandibular inflammation-induced orofacial persistent pain. The inflammatory process in TMJs of rats was induced by Freund's Complete Adjuvant (CFA) administration. The activity and expression of MMPs-2 and 9 were assessed by in situ zymography and conventional zymography, respectively. A glial colocalization with the MMPs was performed using immunofluorescence. The results evidenced both short- and long-term alterations on MMP-2 and -9 expression in the limbic structures following CFA-induced temporomandibular inflammation. The gelatinolytic activity was increased in the central amygdala, hippocampus, hypothalamus, ventrolateral periaqueductal gray (vlPAG), superior colliculus, and inferior colliculus. Finally, an increase of colocalization of MMP-2/GFAP and MMP-9/GFAP in CFA-induced inflammation groups was observed when compared with saline groups in the central amygdala and vlPAG. It is possible to suggest that glial activation is partly responsible for the production of gelatinases in the persistent orofacial pain, and it is involved in the initiation and maintenance of this process, indicating that inhibition of MMPs might be pursued as a potential new therapeutic target for TMD.

Activity and expression of enkephalinase and aminopeptidase N in regions of the mesocorticolimbic system are selectively modified by acute ethanol administration.

Opioid peptides play a key role in ethanol reinforcement and alcohol drinking behavior. However, regulation of opioid levels by peptidase-degrading activities in ethanol's actions in brain is still unclear. The aim of this work was to study the acute effects of ethanol (2.5 g/kg) on enkephalinase (NEP) and aminopeptidase N (APN) activities and expression in regions of the mesocorticolimbic system, as well as on corticosterone levels in serum for up to 24 h after administration. Enzymatic activities were measured by fluorometric assays, mRNA's expression by reverse transcriptase polymerase chain reaction (RT-PCR) and corticosterone levels by radioimmunoassay. Acute ethanol administration modified peptidase activity and expression with different kinetics. Ethanol induced a transitory increase and decrease in NEP and APN activities in the frontal cortex (FC) and ventral tegmental area (VTA), whereas only increases in these activities were observed in the nucleus accumbens (NAcc). Ethanol induced an increase in NEP mRNA in the FC and decreases in APN mRNA in the FC and NAcc. In contrast, ethanol produced biphasic effects on both enzymes expression in the VTA. Corticosterone levels were not changed by ethanol. Our results suggest that NEP and APN could play a main role in ethanol reinforcement through regulation of opioid levels in mesolimbic areas.

Acute administration of alcohol modulates pyroglutamyl amino peptidase II activity and mRNA levels in rat limbic regions.

Released TRH is inactivated by an ectopeptidase, pyroglutamyl aminopeptidase II (PPII). PPII expression and activity are stringently regulated in adenohypophysis, and in rat brain, during kindling stimulation that activates TRHergic neurons. To gain further insight into the possible regulation of PPII, we studied the effect of an acute intraperitoneal ethanol administration that affects TRH content and expression. PPII activity was determined by a fluorometric assay and PPII mRNA levels by semi-quantitative RT-PCR. Activity decreased in frontal cortex 1 h after ethanol injection and, after 6 h, in hippocampus, amygdala and n. accumbens. PPII mRNA levels decreased at 30 and 60 min in frontal cortex and n. accumbens while increased at longer times in these regions and, in hippocampus and hypothalamus. NMDA and GABA(A) receptors' agonists and antagonists were tested at 1 h (+/-ethanol) on PPII activity and mRNA levels, as well as on TRH content and its mRNA. In n. accumbens, PPII mRNA levels decreased by ethanol, MK-801, and muscimol while picrotoxin or NMDA reversed ethanol's inhibition. Ethanol decreased TRH content and increased TRH mRNA levels as MK-801 or muscimol did (NMDA or picrotoxin reverted the effect of ethanol). In frontal cortex, PPII activity was inhibited by ethanol, NMDA and MK-801 with ethanol; its mRNA levels were reduced by ethanol, MK-801 and muscimol (NMDA and picrotoxin reverted ethanol's inhibition). These results show that PPII expression and activity can be regulated in conditions where TRHergic neurons are modulated. Effects of ethanol on PPII mRNA levels as well as those of TRH and its mRNA may involve GABA or NMDA receptors in n. accumbens. Changes observed in frontal cortex suggest combined effects with stress. The response was region-specific in magnitude, tendency and kinetics. These results give further support for brain PPII regulation in conditions that modulate the activity of TRHergic neurons.

Mitogen-activated protein kinase is increased in the limbic structures of the rat brain during the early stages of status epilepticus.

Systemic administration of pilocarpine (PILO) in adult rat produces acute limbic seizures leading to status epilepticus. Recent studies have shown the activation of mitogen-activated protein kinase (MAPK) cascades during experimentally induced seizures. MAPK activation may be triggered by glutamatergic stimulation and may play a key role in signal transduction pathways. In the present study, immunocytochemistry was used to analyze the spatiotemporal distribution pattern of the MAPK protein and its active form (A-MAPK) following PILO-induced status epilepticus. MAPK and A-MAPK immunoreactivities exhibited different patterns of distribution in the brain of normal and epileptic rats. The saline-treated rats, as well as the animals that received PILO but did not evolve to status epilepticus, showed a weak but selective MAPK immunoreactivity, detected in the hippocampal pyramidal neurons, dentate gyrus, hilus, CA3, CA1, and entorhinal, piriform, and cingulate cortices. A-MAPK immunoreactivity was instead observed only in neurites of the CA3 and hilus and in cells of the entorhinal and piriform cortices. In PILO-treated rats, between 30 and 60 min after status epilepticus there was an increase of the immunoreactivity to both antibodies, which were differently distributed throughout several structures of the limbic system. The immunostaining showed a slight decrease after 5 h of status epilepticus. However, MAPK and A-MAPK immunopositivities decreased markedly after 12 h of status epilepticus, returning almost to the basal expression. These findings are consistent with a spatial and time-dependent MAPK expression in selected limbic structures, and its activation could represent an initial trigger for neuronal modifications that may take part in the mechanism underlying acute epileptogenesis and in long-lasting neuropathological changes of the PILO model of epilepsy.