the internal globus pallidus ( gpi ) and substantia nigra pars reticulata ( snr ) are the basal ganglia ( bg ) output nuclei . besides projecting to the thalamus to form the cortico - bg loops ( chevalier et al . , 1985 ;
smith and bolam , 1989 ; alexander and crutcher , 1990 ; delong , 1990 ; smith et al .
, 1998 ; haber , 2003 ) , output nuclei also project to pons and brain stem to control descending pathways and central patterns generators ( cpgs ) that regulate muscular tone and automatic or rhythmic motor responses ( takakusaki et al . , 2003 , 2004 ; grillner et al . , 2008 ) .
in birds , reptiles , and lower vertebrates in which the cortex is not well developed , the control of brain stem nuclei is a main function of the bg ( reiner et al . , 1998 ; grillner et al . ,
2005 , 2008 ; gale and perkel , 2010 ) . in the snr , inhibitory postsynaptic currents ( ipscs )
are in part provided by striatonigral direct pathway terminals ( grofov and rinvik , 1970 ; chevalier et al . , 1985 ; smith and bolam , 1991 ;
, 1996 ; matuszewich and yamamoto , 1999 ) , which possess functional presynaptic dopamine d1-receptors whose activation increases direct pathway inhibition ( porceddu et al .
, 1986 ; altar and hauser , 1987 ; floran et al . , 1990 ; radnikow and misgeld , 1998 ; chuhma et al . ,
enhancement of direct pathway inhibition facilitates movements while its reduction represses them ( albin et al . , 1989 ;
in contrast , subthalamonigral afferents compose the last step of the indirect pathway ( nakanishi et al .
presynaptic modulation of subthalamonigral terminals by dopamine uses both classes of dopamine receptors : d1 and d2 ( ibez - sandoval et al . , 2006 ) .
activation of d1 enhances while activation of d2 depresses subthalamonigral excitatory postsynaptic currents ( epscs ) .
interestingly , simultaneous blockade of both receptors induced larger evoked epscs , suggesting that d2-receptors have more influence than d1-receptors in the modulation of transmission ( ibez - sandoval et al . , 2006 ) .
in parallel , the external globus pallidus ( gpe ) also sends an inhibitory input to snr ( bevan et al . , 1996 ; kita , 2007 ; connelly et al . , 2010 ) .
we found that d2-class receptors regulate these terminals with no sign of d1-receptor modulation as compared to striatonigral d1-mediated modulation in the same preparation .
interestingly , and as shown before for subthalamonigral afferents ( ibez - sandoval et al . , 2006 ) , we found that addition of selective receptor antagonists disclose a tonic action of ambient dopamine levels on both , striatonigral and pallidonigral afferents , supporting the notion that dopamine receptors are sensing extracellular dopamine continuously ( yanovsky et al . , 2003 ) .
furthermore , the blockade of all dopamine receptor types altogether induced a recurrent bursting firing pattern in snr projection neurons , a mode of firing typically seen in both parkinsonian humans and animals ( e.g. , magill et al . , 2001 ; walters et al .
, 2007 ; walters and bergstrom , 2009 ; zold et al . , 2009 ) .
this finding indicates that it is enough to acutely block dopamine receptors to induce a parkinsonian firing pattern in snr neurons .
given the direct projection of snr to brain stem nuclei controlling muscular tone and cpgs activation , a question as to whether dopamine dysfunction in the output nuclei is enough to produce some parkinsonian signs ( morris et al .
, 1994 ; hemsley and crocker , 1998 ; hikosaka et al . , 2000 ;
procedures were carried out in accordance with the national institutes of health guide for care and use of laboratory animals ( nih publications no .
8023 , revised 1996 ) and were approved by the institutional animal care committee of unam .
methods have been reported elsewhere ( beurrier et al . , 2006 ; ibez - sandoval et al . , 2006 ) .
briefly , wistar rats ( 1540 postnatal day ) , were anesthetized with isoflurane , decapitated , and their brains removed .
parasagittal or parahorizontal slices ( 300  m ) containing the neostriatum ( nst ) , globus pallidus ( gp ) , and substantia nigra pars reticulata ( snr ) were obtained with a vibratome in saline of the following composition ( in millimolar ) : 124 choline chloride , 2.5 kcl , 1.0 mgcl2 , 1.2 nah2po4 , 2.0 cacl2 , and 10 glucose ( ~4c 95% o2 , 5% co2 ) .
whole - cell patch - clamp recordings were performed on rat snr neurons ( ibez - sandoval et al . , 2006 , 2007 ) .
neurons within the snr were visualized with infrared differential interference videomicroscopy using a x60 water  immersion objective . for voltage - clamp recordings micropipettes
25  m resistance were filled with internal saline containing high cl ( in millimolar ) : 70 kh2po4 , 36 kcl , 2 mgcl2 , 10 hepes , 1.1 egta , 0.2 na2atp , 0.2 na3gtp , 5  mm qx-314 , 5  mm cscl , and 0.1% biocytin ( ph 7.2 ; 275 mosm / l ) that allowed to record inward ipscs from snr neurons after field stimulation in the internal capsule ( ic ) 0.51.0  mm outside the snr border ( radnikow and misgeld , 1998 ; wallmichrath and szabo , 2002 ) .
bipolar pencil shaped concentric tungsten electrodes , 11.5  m at the tip , and 1  k dc resistance were used . for current - clamp recordings we used internal saline of the following composition ( in millimolar ) : 120 kso3ch4 , 10 nacl , 10 egta , 10 hepes , 1 cacl2 , 2 mgcl2 , 2 atp - mg , 0.3 gtp - na ( ph 7.3 , 290 mosm / l .
superfusion saline contained antagonists for glutamatergic receptors : 10  m 6-cyano-7-nitroquinoxaline-2,3-dione ( cnqx ) and 50  m d-()-2-amino-5-phosphopentanoic acid ( apv ) to isolate ipscs . in parasagittal slices ,
3 out of 10 recordings evoked pallidonigral ipscs and the rest evoked striatonigral ipscs ( figure 1 ) .
in parahorizontal slices 6 out of 10 recordings evoked pallidonigral ipscs and the rest evoked striatonigral ipscs .
ipscs from each source were easily discernible with electrophysiological techniques ( figure 1 ; connelly et al .
, 2010 ) so that when an obvious mixture of ipscs from both sources was obtained , it was discarded from the present analysis .
a paired - pulse protocol was employed with inter - pulse intervals of 50  ms to evaluate changes in the paired - pulse ratio ( ppr ) of evoked ipscs ( ppr  =  2nd ipsc/1st ipsc ) to verify presynaptic actions of transmitters ( ibez - sandoval et al . , 2006 ) .
because striatonigral fibers pass through the gpe , d2-class selective agonists were tested in slices taken from animals with a stereotaxic lesion ( ibotenic acid ) of the gpe ( 1.4  mm ap , 3.4 l , and 4.7  mm v ) and compared to recordings obtained without a lesion .
ibotenic acid solution ( dissolved in pbs adjusted to ph 7.4 with naoh 3.0  g/0.4  l ) was used to lesion the gpe .
( a ) striatonigral inhibitory postsynaptic currents ( ipscs ) exhibited paired - pulse facilitation .
( c ) short - term synaptic plasticity ( stp ) from striatonigral afferents is facilitation .
( e ) intensity  amplitude plots ( i  a plots ) from striatonigral ipscs exhibit a sigmoidal shape .
( f ) i  a plots from pallidonigral ipscs exhibit a jump to maximal amplitude after reaching threshold .
a plots were fitted to : a(i )  =  amax/(1  +  e ) where a(i )  =  ipsc amplitude as a function of stimulus intensity , amax  =  maximal amplitude reached , k  =  slope factor , and ih  =  stimulus intensity necessary to reach ipsc amplitude equal to half maximal amplitude .
drugs were stored as dry aliquots and stock solutions were prepared prior to each experiment and added to the perfusion solution in the final concentration indicated .
skf 81297 : ( )-6-chloro-2,3,4,5-tetrahydro-1-phenyl-1h-3-benzazepine hydrobromide ( 10  nm50  m ) ; sch 23390 : 7-chloro-8-hydroxy-3-methyl-1-phenyl-2,3,4,5-tetrahydro-1h-3-benzazepine ( 50  nm and 1  m ) ; cnqx : 6-cyano-7-nitroquinoxaline-2,3-dione ( 10  m ) , d - ap-5  : d-()-2-amino-5-phosphopentanoic acid ( 50  m ) , bicuculline methiodide or methchloride ( 10  m ) , and qx-314 ( 5  mm inside the recording pipette ) , all were obtained from sigma - aldrich ( st . louis mo , usa ) .
the neurons in the present work were snr projection neurons ( n  =  250 ; see : ibez - sandoval et al . , 2006 ) .
ipscs parameters from different afferents , i.e. , striatonigral and pallidonigral were compared with mann 
ipscs parameters in the same synapses before and after adding a given drug were compared with wilcoxon 's t - tests .
at least 10  min of stable recordings before and after administering the drugs were used to reach stable average ipscs amplitudes .
differences in parameters of fitted functions were compared by using their estimation error and student 's t - tests .
figure 1 shows that striatonigral and pallidonigral synaptic inputs can not be confused ( connelly et al . , 2010 ) .
for similar stimulus , evoked striatonigral ipscs were significantly smaller than pallidonigral ipscs ( mean    sem for the first ipsc of a paired response ) : 235    50  pa ( n  =  32 ) vs. 1399    152  pa ( n  =  20 ; p  <  0.0001 ) , respectively .
responses to paired - pulse stimulation were also different : striatonigral ipscs showed paired - pulse facilitation : ppr  =  1.41    0.07 ( figures 1a , g ; n  =  30 ) whereas pallidonigral ipscs always exhibited paired - pulse depression : ppr  =  0.4    0.06 ( figures 1b , g ; n  =  19 ; p  <  0.0001 ) , although not always as strong as that depicted in figure 1d .
short - term plasticity ( 10 pulses at 20  hz ) exhibited facilitation without failures in striatonigral connections ( figure 1c ; n  =  8) , whereas it exhibited depression with intermittent failures in pallidonigral synapses ( figure 1d ; n  =  4 ) .
the decay time constant / rise time ratio was also different : 4.5    0.5 ( n  =  30 ) in striatonigral synapses and 12.6    0.8 in pallidonigral synapses ( n  =  19 ; p  <  0.0001 ) showing that pallidonigral ipscs are briefer than striatonigral ones ( connelly et al . , 2010 ) . intensity  amplitude ( i  a ) relationships were also constructed and fitted to a three parameter sigmoidal function ( tecuapetla et al . , 2005 ) : where a(i )  =  ipsc amplitude as a function of stimulus intensity , amax  =  maximal amplitude reached , k  =  slope factor , and ih  =  stimulus intensity necessary to reach ipsc amplitude equal to half maximal amplitude .
all three parameters were significantly different when comparing ipscs from striatonigral vs. pallidonigral afferents : amax : 430    3  pa vs. 1512    10  pa ( n  =  8 ; p  < 
0.0001 ) ; k  =  3.6    0.3 vs. 14    5 (
n  =  8 ; p  <  0.0001 ) and ih : 2.3    0.2 vs. 1.1    0.1 ( n  =  8 ; p  <  0.005 ) .
these features coincide with a previous report ( connelly et al . , 2010 ) and were verified qualitatively by evoking ipscs from either the subthalamic nucleus ( nst ) or the gpe , however , in these occasions ipscs were considerably smaller and amplitude could not be compared by itself given the different distances from stimulating and recording electrodes ( not shown ) .
striatonigral ipscs features have been well described ( radnikow and misgeld , 1998 ; wallmichrath and szabo , 2002 ; beurrier et al . , 2006 ; chuhma et al . ,
2011 ) as well as ipscs from pallidal origin making synapses into the snr and other nuclei ( tecuapetla et al .
, 2005 ; baufreton et al . , 2009 ; connelly et al . , 2010 ) .
finally , three of these variables were used to build a cluster plot ( figure 1 g ) which confirmed the easiness to separate ipscs from each source .
as it has been repeatedly demonstrated , the action of dopaminergic d1-class selective agonists at nanomolar concentrations was that of enhancing striatonigral ipscs ( floran et al .
, 1990 ; radnikow and misgeld , 1998 ; chuhma et al . , 2011 )
: striatonigral ipsc increased 153    10% after 300  nm skf 81297 ( n  =  15 ; p  <  0.001 ) and the paired - pulse ratio ( ppr  =  ipsc2/ipsc1 ) decreased from 1.4    0.13 in the control to 1.0    0.12 during skf 81297 ( p  <  0.001 ) , confirming a presynaptic modulation .
these actions were reversible and blocked by 100  nm of the d1-antagonist sch 23390 ( n  =  5 ; not shown here but see below ) indicating that at these concentrations the action is specific .
in addition , here we show that the agonists have no significant action on pallidonigral ipscs ( cf .
, figures 2a , b ) . contrasting actions of d1- and d2-receptor agonists on striatonigral and pallidonigral ipscs . ( a ) 100  nm of the selective d1-class receptor agonist , skf 81297 , enhanced striatonigral ipscs . ( b )
( c ) 500  nm of the selective d2-class receptor agonist , quinelorane , had no significant action on striatonigral ipscs .
record 3 in each frame is the superimposition of records 1 and 2 after normalization of the first ipsc to better appreciate the ppr change when it is present .
contrasting results were obtained when a selective d2-class receptor agonist , 500  nm quinelorane , was employed : striatonigral ipscs suffered no significant alteration ( figure 2c ; n  =  5 ns ; chuhma et al . , 2011 ) whereas pallidonigral ipscs were significantly inhibited ( figure 2d ) by 73    18% ( figure 2d ; n  =  19 ; p  <  0.0001 ) .
ppr increased from 0.76    0.1 to 1.83    0.2 ( p  <  0.001 ) , suggesting a presynaptic action .
these actions were reversible and blocked by 500  nm sulpiride , a d2-antagonist ( n  =  5 ; not shown but see below ) . summarizing , physiological action of d1-receptor agonists on striatonigral terminals is that of ipsc enhancement as previously shown ( floran et al . , 1990 ; radnikow and misgeld , 1998 ; chuhma et al . , 2011 ) with no action on pallidonigral terminals
on the other hand , physiological action of d2-receptor agonists on pallidonigral ipscs is that of depression with no significant action on striatonigral terminals .
therefore we were forced to infer that reports about d1-mediated inhibition of striatonigral terminals ( miyazaki and lacey , 1998 ) were either involving a non - specific action , a pallidal contamination , or both . to test this hypothesis we used larger micromolar concentrations of the d1-agonist while evoking ipscs from both pathways .
figures 3a , b show that 5  m skf 81297 decreased ipscs evoked from both set of terminals .
striatonigral ipsc decreased 82    13% ( n  =  18 ; p  <  0.001 ) and pallidonigral responses decreased by 35    15% ( n  =  6 ; p  <  0.005 ) .
these actions could not be blocked by micromolar concentrations of sch 23390 ( not shown ) , suggesting that they were not specific . in view of these results
response relationship ( c  r plot ) using a wide range of skf 81297 concentrations while stimulating striatonigral afferents .
this c  r plot can be seen in figure 3c : it is biphasic .
when the hill equation was fitted to the ascending ( specific part ) ec50 was 440    60  nm and the hill coefficient 1.6    0.2 , suggesting cooperativity and a specific action at submicromolar concentrations .
moreover , the fact that pallidonigral inputs are also affected when they do not respond when submicromolar concentrations of agonists are used confirmed non - specific actions .
record 3 in each frame is the superimposition of representative records 1 and 2 after normalization of the first ipsc to better appreciate the ppr change when it is present .
( c ) concentration  response relationship ( c  r plot ) showed a biphasic response on striatonigral responses : once 3  m are surpassed , ipscs decay with increasing concentrations .
ascending c  r plot was fitted with the hill equation finding a submicromolar ec50 .
given the low concentrations of agonists needed to activate d1- and d2-class receptors in their respective terminals ( striatonigral and pallidonigral ) we inferred that , perhaps , endogenous extracellular dopamine exerts a tonic action on these receptors .
figures 4a , b show that 50  nm of a d1-class receptor selective antagonist , sch 23390 , are enough to inhibit striatonigral ipscs with no significative action on pallidonigral ipsc .
striatonigral ipsc decreased from 319    75 in the control to 150    11  pa after 50  nm sch 23390 ( n  =  12 ; p  <  0.02 )
. figures 4c , d confirm these findings and further show that potency and speediness of d1-action is concentration dependent .
almost abolished  when a low micromolar antagonist concentration is maintained in the superfusion ( figure 4c ) .
traces chosen at different times during the time course , superimposed , and normalized to the amplitude of the first ipsc , show that the ppr is greatly increased from 1.9    0.4 in the control to 2.4    0.6 after sch 23390 ( n  =  21 ; p  <  0.001 ; when the ipsc is abolished ppr can not be measured ) ; confirming a presynaptic site of action .
on the other hand , sch 23390 did not produce any action on pallidonigral ipsc at any concentration ( figure 4d ) .
in summary , d1-receptors in direct pathway striatonigral terminals are sensitive detectors of extracellular dopamine . moreover ,
( a ) 50  nm of selective dopamine d1-receptor antagonist , sch 3390 , are enough to reduce striatonigral ipsc amplitude .
( c ) increasing d1-antagonist concentration ( 1  m ) greatly reduced ipsc while speeding up dopamine actions on striatonigral ipsc , suggesting a concentration dependent mechanism .
( e ) 500  nm u-9914a , a selective d3-type receptor antagonist , significantly enhanced pallidonigral ipsc .
( f ) 500  nm ly750 , a selective d4-type receptor antagonist , significantly increased pallidonigral ipsc .
record 3 in each frame is the superimposition of representative records 1 and 2 after normalization of the first ipsc to better appreciate the ppr change when it is present .
in addition , figures 4e , f illustrate the actions of selective antagonists for d3- and d4-type dopamine receptors : 500  nm u-9914a , a selective d3-type receptor antagonist enhanced pallidonigral ipscs by 262    16% after ( n  =  12 ; p  < 
0.001 ) while the ppr decreased from 0.87    0.08 in the control to 0.51    0.1 during the antagonist . on the other hand , 500  nm ly750 , a selective d4-type receptor antagonist increased pallidonigral ipscs by 170    20% ( n  =  7 ; p  <  0.001 ) while ppr decreased from 0.78    0.07 in the control to 0.57    0.09 during the blockade .
sulpiride a generic d2-class receptor antagonist had similar actions ( not shown ) : pallidonigral ipscs increased by 150    14% ( n  =  3 ) while ppr decreased from 0.77    0.03 in the control to 0.35    0.01 during blockade . to summarize , d3/4-receptors in pallidonigral terminals ( murray et al . , 1994 ; bevan et al . , 1996 ; marshall et al . , 2001
, 2009 ; gasca - martinez et al . , 2010 ) are sensitive to extracellular dopamine , which has the role of tonically repressing the force of these synapses .
some functional differences in the actions of these receptor types perhaps deserve further investigation ( cf .
, figures 4e , f ) . finally , because subthalamonigral terminals are also tonically controlled by presynaptic dopamine receptors ( d1- and d2-class ; ibez - sandoval et al . , 2006 ) and because blockade of these receptors enhance subthalamonigral epscs ( ibez - sandoval et al . , 2006 ) , we propose the following hypothesis based in the present and previous results ( radnikow and misgeld , 1998 ; acosta - garca et al . , 2009 ; chuhma et al . , 2011 ) : that acute blockade of both d1- and d2-class ( including d3/4-types ) receptors ( by 1  m sch 23390 plus 1  m sulpiride ) altogether may decrease direct pathway synapses ( striatonigral ) and , at the same time , enhance indirect pathway synapses ( subthalamonigral and pallidonigral ) , both actions being required in physiopathological models of parkinsonism to elicit the signs of the disease . the result can be seen in figure 5 : snr neurons shifted from a tonic firing pattern in the control to a bursting firing pattern after blockade of dopamine receptors ( n  =  7 neurons ; ibez - sandoval et al . , 2007 ) .
spontaneous tonic firing was the usual firing pattern recorded in snr neurons in control conditions ( top ) . during acute blockade of dopamine d1- and d2-class receptors
( 1  m sch 23390 plus 1  m sulpiride ) snr firing pattern shifted from tonic to a bursting pattern in n  =  7 cells ; two cells exhibited bursting before adding the blockers . to see whether besides striatonigral and pallidonigral afferents
, subthalamonigral afferents were also contributing to this firing mode , the glutamate nmda - receptor antagonist , 50  m apv was added to the bath saline ( bottom ) .
the firing tended to return to a tonic firing pattern but it was frequently interrupted by sudden hyperpolarizations .
one of the most supported neuronal correlates of parkinsonism : recurrent bursting in snr neurons , was similar to that previously reported in vivo and in vitro ( hammond et al .
, 2007 ; ibez - sandoval et al . , 2007 ; walters et al . , 2007 ; walters and bergstrom , 2009 ; zold et al . , 2009 )
transitions from tonic to bursting firing mode can rarely be seen spontaneously in control preparations ( ibez - sandoval et al . , 2007 ) . to see whether excitatory subthalamonigral afferents ,
that is , the stn - gp circuit , was participating in this firing behavior , we added the nmda - receptor antagonist apv ( 50  m ) to provoke a partial block of stn influence .
the result was that the firing pattern became less bursty and more tonic ( ibez - sandoval et al . , 2007 ) , however , firing was still abruptly interrupted by sudden hyperpolarizations , probably coming from enhanced pallidonigral inputs .
the present work shows that extracellular dopamine concentrations are tonically being sensed by the synaptic terminals of inhibitory inputs to the snr in opposite ways .
thus , blockade of d1-type receptors in striatonigral ( direct pathway ) afferents decreased striatonigral inhibition while blockade of d3/4-types receptors in pallidonigral terminals enhanced pallidonigral inhibition .
still other inputs to the snr have to be studied to see whether they are presynaptically modulated .
in addition , they exhibit short - term facilitation and are positively regulated by d1-receptors ( floran et al . , 1990 ; radnikow and misgeld , 1998 ; connelly et al . , 2010 )
strong evidence for the last property has been obtained with optogenetic techniques ( chuhma et al . , 2011 ) , supporting the present and previous reports . in comparison ,
pallidonigral ipscs are larger and exhibit different degrees of short - term depression ( connelly et al . , 2010 ) .
notably , short - term facilitation works as a high pass filter of incoming inputs whereas short - term depression works as a low pass filter ( abbott and regehr , 2004 ) .
in contrast , subthalamonigral excitatory synapses are controlled by both d1- and d2-class receptors ( ibez - sandoval et al . ,
selective dopamine receptor agonists exert their specific actions at nanomolar or low micromolar concentrations but higher concentrations become non - specific , thus explaining previous contradictions ( cf . , miyazaki and lacey , 1998 ; radnikow and misgeld , 1998 ) and probably similar discrepancies within the striatal / accumbinal circuitry ( guzman et al . , 2003 ; taverna et al . ,
2005 ) . to summarize : d1-receptor activation enhances gaba release in terminals from medium spiny neurons of the direct pathway whereas d3/4-receptor activation represses gaba release from pallidonigral terminals .
because nanomolar concentrations of selective receptor agonist are needed to modify transmission , we hypothesized that administrations of selective dopamine receptor antagonist would disclose the actions of endogenous dopamine present in the extracellular space .
therefore , to disclose the action of ambient endogenous dopamine we applied selective d1- and d3/4-receptor antagonists .
our findings were that according to the concentration , suppression of endogenous dopamine action greatly reduced striatonigral transmission while it enhanced pallidonigral transmission .
these results indicated that extracellular dopamine concentrations are regulating synaptic release probability in both types of connections , increasing release probability in striatonigral synapses and decreasing release probability in pallidonigral synapses .
therefore , the following working hypothesis resulted from the present experiments : tonic levels of dopamine in snr are necessary to maintain the normal function of direct pathway connections and maintain in check indirect pathway synapses from both pallidonigral terminals ( present work ) and subthalamonigral terminals ( ibez - sandoval et al . , 2007 )
. the logical functional consequence of this hypothesis was tested : blockade of tonic dopamine action would enhance indirect pathway transmission and reduce direct pathway input thus yielding a previously observed neuronal correlate of parkinsonism : snr neurons shifted their tonic firing pattern to a bursting firing pattern typical of subjects with the disease ( takakusaki et al . , 2004 ; rivlin - etzion et al . , 2006 ;
hammond et al . , 2007 ; ibez - sandoval et al . , 2007 ; walters et al .
tonic spontaneous firing is preserved in the slice preparation in both gpe and stn ( e.g. , beurrier et al . , 1999 ; chan et al . , 2011 ) .
calcium imaging techniques recording dozens of cells simultaneously within the striatal circuit show that there is always some spontaneous activity in striatal spiny neurons in control conditions ( carrillo - reid et al . , 2008 ) .
moreover , there is a great amount of convergence between the striatum and the substantia nigra and inhibitory striatonigral events are of large amplitude ( e.g. , chuhma et al . , 2011 ) .
therefore , the lost balance between direct and indirect pathways by the acute blockade of dopamine receptors is the most probable cause of bursting in snr neurons during the present experiments .
postsynaptic dopamine receptors on snr neurons can not explain bursting because their function is to increase tonic firing frequency via a cation current ; their blockade resulting in lower tonic frequency with irregularities , but not continuous bursting behavior ( lee and tepper , 2007 ; zhou et al . , 2009 ) .
because the change in firing pattern of snr neurons was achieved acutely by blocking dopamine receptors one can speculate what would happen during a chronic absence of dopamine in the output nuclei of the bg .
the absence of dopamine ( e.g. , parkinsonism ) may reduce the function of direct pathway synapses in such a way that maintaining this state of affairs in the long - time would lead to loss of direct pathway synapses due to long - term synaptic plasticity .
in contrast , maintaining a high function in pallidonigral and subthalamonigral synapses would produce long - term potentiation of these synapses .
these events taken together may tend to change the circuitry permanently , making not only l - dopa inefficient but , perhaps , making the chronic diseased circuitry radically different than the control or healthy circuit : that is , more dependent on the subthalamopallidal loop ( magill et al .
bursting in snr neurons leads to tremor and rigidity ( hemsley and crocker , 1998 ; takakusaki et al . ,
moreover , this configuration of synaptic weights would lead to the loss of high pass filtering of snr inputs ( direct pathway 's short - term facilitation ) and an increase in low pass filtering of snr inputs ( pallidonigral short - term depression ; abbott and regehr , 2004 ) setting the stage to favor the entrance to an akinetic frequency lock ( hutchison et al .
, 2004 ; avila et al . , 2010 ) . consequently , therapeutic ways of activating the direct pathway in the chronic patient may be fundamental to avoid irreversible plastic changes in the network ( hammond et al .
, 2007 ; walters and bergstrom , 2009 ; zold et al . , 2009 ;
, they exhibit short - term facilitation and are positively regulated by d1-receptors ( floran et al . , 1990 ; radnikow and misgeld , 1998 ; connelly et al . , 2010 )
strong evidence for the last property has been obtained with optogenetic techniques ( chuhma et al . , 2011 ) , supporting the present and previous reports . in comparison ,
pallidonigral ipscs are larger and exhibit different degrees of short - term depression ( connelly et al . , 2010 ) .
notably , short - term facilitation works as a high pass filter of incoming inputs whereas short - term depression works as a low pass filter ( abbott and regehr , 2004 ) .
in contrast , subthalamonigral excitatory synapses are controlled by both d1- and d2-class receptors ( ibez - sandoval et al . , 2006 ) .
selective dopamine receptor agonists exert their specific actions at nanomolar or low micromolar concentrations but higher concentrations become non - specific , thus explaining previous contradictions ( cf . , miyazaki and lacey , 1998 ; radnikow and misgeld , 1998 ) and probably similar discrepancies within the striatal / accumbinal circuitry ( guzman et al . , 2003 ; taverna et al . ,
2005 ) . to summarize : d1-receptor activation enhances gaba release in terminals from medium spiny neurons of the direct pathway whereas d3/4-receptor activation represses gaba release from pallidonigral terminals
because nanomolar concentrations of selective receptor agonist are needed to modify transmission , we hypothesized that administrations of selective dopamine receptor antagonist would disclose the actions of endogenous dopamine present in the extracellular space .
therefore , to disclose the action of ambient endogenous dopamine we applied selective d1- and d3/4-receptor antagonists .
our findings were that according to the concentration , suppression of endogenous dopamine action greatly reduced striatonigral transmission while it enhanced pallidonigral transmission .
these results indicated that extracellular dopamine concentrations are regulating synaptic release probability in both types of connections , increasing release probability in striatonigral synapses and decreasing release probability in pallidonigral synapses .
therefore , the following working hypothesis resulted from the present experiments : tonic levels of dopamine in snr are necessary to maintain the normal function of direct pathway connections and maintain in check indirect pathway synapses from both pallidonigral terminals ( present work ) and subthalamonigral terminals ( ibez - sandoval et al . , 2007 ) .
the logical functional consequence of this hypothesis was tested : blockade of tonic dopamine action would enhance indirect pathway transmission and reduce direct pathway input thus yielding a previously observed neuronal correlate of parkinsonism : snr neurons shifted their tonic firing pattern to a bursting firing pattern typical of subjects with the disease ( takakusaki et al . , 2004 ; rivlin - etzion et al . , 2006 ;
tonic spontaneous firing is preserved in the slice preparation in both gpe and stn ( e.g. , beurrier et al . , 1999 ; chan et al . , 2011 ) .
calcium imaging techniques recording dozens of cells simultaneously within the striatal circuit show that there is always some spontaneous activity in striatal spiny neurons in control conditions ( carrillo - reid et al . , 2008 ) .
moreover , there is a great amount of convergence between the striatum and the substantia nigra and inhibitory striatonigral events are of large amplitude ( e.g. , chuhma et al . , 2011 ) .
therefore , the lost balance between direct and indirect pathways by the acute blockade of dopamine receptors is the most probable cause of bursting in snr neurons during the present experiments .
postsynaptic dopamine receptors on snr neurons can not explain bursting because their function is to increase tonic firing frequency via a cation current ; their blockade resulting in lower tonic frequency with irregularities , but not continuous bursting behavior ( lee and tepper , 2007 ; zhou et al . , 2009 ) .
because the change in firing pattern of snr neurons was achieved acutely by blocking dopamine receptors one can speculate what would happen during a chronic absence of dopamine in the output nuclei of the bg .
the absence of dopamine ( e.g. , parkinsonism ) may reduce the function of direct pathway synapses in such a way that maintaining this state of affairs in the long - time would lead to loss of direct pathway synapses due to long - term synaptic plasticity . in contrast , maintaining a high function in pallidonigral and subthalamonigral synapses would produce long - term potentiation of these synapses .
these events taken together may tend to change the circuitry permanently , making not only l - dopa inefficient but , perhaps , making the chronic diseased circuitry radically different than the control or healthy circuit : that is , more dependent on the subthalamopallidal loop ( magill et al .
bursting in snr neurons leads to tremor and rigidity ( hemsley and crocker , 1998 ; takakusaki et al . ,
moreover , this configuration of synaptic weights would lead to the loss of high pass filtering of snr inputs ( direct pathway 's short - term facilitation ) and an increase in low pass filtering of snr inputs ( pallidonigral short - term depression ; abbott and regehr , 2004 ) setting the stage to favor the entrance to an akinetic frequency lock ( hutchison et al .
, 2004 ; avila et al . , 2010 ) . consequently , therapeutic ways of activating the direct pathway in the chronic patient may be fundamental to avoid irreversible plastic changes in the network ( hammond et al .
, 2007 ; walters et al . , 2007 ; walters and bergstrom , 2009 ; zold et al . , 2009 ; bateup et al . , 2010 ; kravitz et al . , 2010 )
the authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest .
 previous work has shown the functions associated with activation of dopamine presynaptic receptors in some substantia nigra pars reticulata ( snr ) afferents : ( i ) striatonigral terminals ( direct pathway ) posses presynaptic dopamine d1-class receptors whose action is to enhance inhibitory postsynaptic currents ( ipscs ) and gaba transmission . 
 ( ii ) subthalamonigral terminals posses d1- and d2-class receptors where d1-class receptor activation enhances and d2-class receptor activation decreases excitatory postsynaptic currents . here 
 we report that pallidonigral afferents posses d2-class receptors ( d3 and d4 types ) that decrease inhibitory synaptic transmission via presynaptic modulation . 
 no action of d1-class agonists was found on pallidonigral synapses . 
 in contrast , administration of d1-receptor antagonists greatly decreased striatonigral ipscs in the same preparation , suggesting that tonic dopamine levels help in maintaining the function of the striatonigral ( direct ) pathway . 
 when both d3 and d4 type receptors were blocked , pallidonigral ipscs increased in amplitude while striatonigral connections had no significant change , suggesting that tonic dopamine levels are repressing a powerful inhibition conveyed by pallidonigral synapses ( a branch of the indirect pathway ) . 
 we then blocked both d1- and d2-class receptors to acutely decrease direct pathway ( striatonigral ) and enhance indirect pathways ( subthalamonigral and pallidonigral ) synaptic force . 
 the result was that most snr projection neurons entered a recurrent bursting firing mode similar to that observed during parkinsonism in both patients and animal models . 
 these results raise the question as to whether the lack of dopamine in basal ganglia output nuclei is enough to generate some pathological signs of parkinsonism . 
Introduction
Materials and Methods
Results
Discussion
Striatonigral and pallidonigral IPSCs are different
Striatonigral and pallidonigral dopamine receptors sense extracellular ambient endogenous dopamine
Functional consequences
Conflict of Interest Statement