Nigral Degeneration with Inclusion Body Formation and Behavioral Changes in Rats after Proteasomal Inhibition

Abstract

Objective: We were interested in studying nigral degeneration with inclusion body formation and behavioral changes in rats after proteasomal inhibition. Methods: Observation of progressive behavioral and pathological changes in rats following a unilateral nigral injection of lactacystin, a selective proteasome inhibitor. Results: After administration at a concentration of 10 μg (2 μl) of lactacystin, when tyrosine hydroxylase (TH) immunostaining decreased gradually in the substantia nigra pars compacta (SNc) and corpus striatum, α-synuclein-immunopositive inclusion appeared extensively in the surviving neurons. We also observed the degeneration of diverse cellular organelles by transmission electron microscopy. The effect of cellular organelle degeneration on behavior, a clinical index, was striking and was statistically significant. Over the 3 weeks following the administration of lactacystin, a highly significant decrease in TH immunostaining was observed and α-synuclein-immunopositive inclusions gradually appeared. Interestingly, there was a strong correlation in behavioral changes and the increase in α-synuclein-immunopositive inclusions whereas the decrease in TH immunostaining did not seem to induce any behavioral changes. Conclusions: Our results reveal that unilateral nigral proteasome inhibition induces degeneration in the SNc and corpus striatum as well as behavioral changes demonstrating strong time dependence. Behavioral changes were driven by the formation of α-synuclein inclusions, but not by decreased TH neurons.

The Discovery of a Highly Selective 5,6,7,8‐Tetrahydrobenzo[4,5]thieno[2,3‐d]pyrimidin‐4(3H)‐one SIRT2 Inhibitor that is Neuroprotective in an in vitro Parkinson’s Disease Model

“…[74,75] This translates to the formation of ubiquitin/aSyn immunopositive inclusions similar to those found in Parkinson's disease and in lactacystin-treated dopaminergic neurons, and subsequent neurodegeneration, both in vitro [75] and in vivo. [74,[76][77][78][79] We therefore used lactacystin to model Parkinsonian neuronal cell death in the N27 mescencephalic dopaminergic cell line (Figure 8). [80] In line with previous findings using a SIRT2 inhibitor, [34] here we observe that cultured N27 cells pretreated for 48 hours with compound 10 and subsequently treated with lactacystin display significantly greater neuronal survival than vehicletreated controls, as ascertained from a combination of cell viability assays ( Figure S6 in the Supporting Information).…”

Decreased behavioral response to intranigrally administered GABAA agonist muscimol in the lactacystin model of Parkinson's disease may result from partial lesion of nigral non-dopamine neurons: Comparison to the classical neurotoxin 6-OHDA

“…Since some previous in vivo studies [18,19,23] have used higher doses of lactacystin to examine motor deficits than this used in our study, we also investigated, using of various parameters of locomotor activity, whether unilateral intranigral injection of 2.5 g/2 l of lactacystin would produce akinesia in rats comparable to akinesia induced by 6-OHDA (8 g/2 l). We demonstrated that 4 weeks after the lesion, the lactacystin group demonstrated a decrease in the total distance traveled and rearing, as well as an increase in the resting time.…”

“…Neuronal degeneration was accompanied by a sharp decrease in the striatal DA content and impairment of the motor dysfunction, reversed by l-DOPA treatment [21]. Furthermore, lactacystin treatment led to the formation of cytoplasmic ␣-synuclein-and ubiquitinimmunopositive inclusion in the SN [18,19,[22][23][24][25]. Thus, the advantage of the lactacystin model over the conventional 6-OHDA model is that, besides the loss of DA neurons in the SN, it represents the cardinal feature of PD, namely abnormal protein degradation.…”

The analysis of differential induction of hsp70 in the related cerebra domains and nerve fibers of rats after proteasome inhibiton

“…In our present study, we have constructed PD model by proteasome inhibitors (lactacystin) injected into striatum and/or SNc as per established protocols. 8 Apart from our model, various other neurotoxin-based models formulated after treating animals via various routes have been developed. But, these models have their own shortcomings such as 1-methyl-4-phenyl- 1,2,3,6-tetrahydropyridine (MPTP) model (no typical Lewy bodies); 12 6-hydroxydopamine (without Lewy bodies); 13 rotenone (variable individual susceptibility) 14 or mouse alpha-synuclein overexpression (no loss of DA neurons in SNc).…”

“…For each animal, an injection cannula (a 30-G stainless-steel cannula connected to a 10μl Hamilton syringe, driven by a microinfusion pump) was slowly inserted through a hole drilled in the skull, into the central part of the left SNc unilaterally using the following coordinates (in mm): anteroposterior (AP), –5.2; mediolateral (ML), ±2.2; dorsoventral (DV), –7.2. 8 Lactacystin, a specific proteasome inhibitor (Sigma-Aldrich Corp, St. Louis, MO, USA), was inserted in left SNc unilaterally ( n = 60, 30 infused with lactacystin, and 30 infused with saline as control), whereas contralateral sides were intact. Infusions were performed during the surgery, when lactacystin, dissolved in physiological saline (10μg/2μl), was delivered at a flow rate of 0.5μl/min for 4 min.…”